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Peptide Synthesis

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Pierce Protein Methods

Page contents

Introduction peptide synthesis
Applications for synthetic peptides

Process of synthesizing peptides

Common protecting scheme-specific solvents
Peptide synthesis strategies
Peptide purification

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Introduction to peptide synthesis

Peptide synthesis is characterized as the formation of a peptide bond between two amino acids. While there is no definitive definition of a peptide, it usually refers to flexible (little secondary structure) chains of up to 30-50 amino acids.

The ability to form peptide bonds to link amino acids together is over 100 years old, although the first peptides to be synthesized, including oxytocin and insulin, did not occur for another 50-60 years, demonstrating the difficult task of chemically synthesizing chains of amino acids (1). In the last 50 years, advances in protein synthesis chemistry and methods have developed to the point where peptide synthesis today is a common approach in even high-throughput biological research and product and drug development (2).

The benefit of peptide synthesis strategies today is that besides having the ability to make peptides that are found in biological specimens, creativity and imagination can be tapped to generate unique peptides to optimize a desired biological response or other result. This page highlights the important aspects of peptide synthesis, the most common methods of synthesis and purification and the strengths and limitations of the respective strategies.

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Applications of synthetic peptides.

The invention of peptide synthesis in the fifties and sixties spurred the development of different application areas in which synthetic peptides are now used, including the development of epitope-specific antibodies against pathogenic proteins, the study of protein functions and the identification and characterization of proteins. Furthermore, synthetic peptides are used to study enzyme-substrate interactions within important enzyme classes such as kinases and proteases, which play a crucial role in cell signaling.

In cell biology, receptor binding or the substrate recognition specificity of newly discovered enzymes can often be studied using sets of homologous synthetic peptides. Synthetic peptides can resemble naturally occurring peptides and act as drugs against cancer and other major diseases. Finally, synthetic peptides are used as standards and reagents in mass spectrometry (MS)-based applications. Synthetic peptides play a central role in MS-based discovery, characterization and quantitation of proteins, especially those that serve as early biomarkers for diseases.

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Peptide synthesis most often occurs by coupling the carboxyl group of the incoming amino acid to the N-terminus of the growing peptide chain. This C-to-N synthesis is opposite from protein biosynthesis, during which the N-terminus of the incoming amino acid is linked to the C-terminus of the protein chain (N-to-C). Due to the complex nature of in vitro protein synthesis, the addition of amino acids to the growing peptide chain occurs in a precise, step-wise and cyclic manner. And while the common methods of peptide synthesis have some critical differences, they all follow the same step-wise method to add amino acids one-at-a-time to the growing peptide chain.

Peptide deprotection

Because amino acids have multiple reactive groups, peptide synthesis must be carefully performed to avoid side reactions that can reduce the length and cause branching of the peptide chain. To facilitate peptide formation with minimal side reactions, chemical groups have been developed that bind to the amino acid reactive groups and block, or protect, the functional group from nonspecific reaction.

Purified, individual amino acids used to synthesize peptides are reacted with these protecting groups prior to synthesis, and then specific protecting groups are removed from the newly added amino acid (a step called deprotection) just after coupling to allow the next incoming amino acid to bind to the growing peptide chain in the proper orientation. Once peptide synthesis is completed, all remaining protecting groups are removed from the nascent peptides. Three types of protecting groups are generally used, depending on the method of peptide synthesis, and are described below.

The amino acid N-termini are protected by groups that are termed "temporary" protecting groups, because they are relatively easily removed to allow peptide bond formation. Two common N-terminal protecting groups are tert-butoxycarbonyl ( Boc ) and 9-fluorenylmethoxycarbonyl ( Fmoc ), and each group has distinct characteristics that determine their use. Boc requires a moderately strong acid such as trifluoracetic acid (TFA) to be removed from the newly added amino acid, while Fmoc is a base-labile protecting group that is removed with a mild base such as piperidine.

Boc chemistry was first described in the 1950s and requires acidic conditions for deprotection, while Fmoc, which was not reported for another twenty years, is cleaved under mild, basic conditions (3,4,5,6). Because of the mild deprotection conditions, Fmoc chemistry is more commonly used in commercial settings because of the higher quality and greater yield, while Boc is preferred for complex peptide synthesis or when non-natural peptides or analogs that are base-sensitive are required.

The use of a C-terminal protecting group depends on the type of peptide synthesis used; while liquid-phase peptide synthesis requires protection of the C-terminus of the first amino acid (C-terminal amino acid), solid-phase peptide synthesis does not, because a solid support (resin) acts as the protecting group for the only C-terminal amino acid that requires protection (see Protein Synthesis Strategies).

Amino acid side chains represent a broad range of functional groups and are therefore a site of considerable side chain reactivity during peptide synthesis. Because of this, many different protecting groups are required, although they are usually based on the benzyl (Bzl) or tert-butyl (tBu) group. The specific protecting groups used during the synthesis of a given peptide vary depending on the peptide sequence and the type of N-terminal protection used (see next paragraph). Side chain protecting groups are known as permanent protecting groups, because they can withstand the multiple cycles of chemical treatment during the synthesis phase and are only removed during treatment with strong acids after synthesis is complete.

Because multiple protecting groups are normally used in peptide synthesis, it is evident that these groups must be compatible to allow deprotection of distinct protecting groups while not affecting other protecting groups. Protecting schemes are therefore established to match protecting groups so that deprotection of one protecting group does not affect the binding of the other groups. Because N-terminal deprotection occurs continuously during peptide synthesis, protecting schemes have been established in which the different types of side chain protecting groups (Bzl or tBu) are matched to either Boc or Fmoc, respectively, for optimized deprotection. These protecting schemes also incorporate each of the steps of synthesis and cleavage, as described in the table and in later sections of this page.

Protecting Scheme	Deprotection	Coupling	Cleavage	Wash
Boc/Bzl	TFA	Coupling agent in DMF	HF, HBr, TFMSA	DMF
Fmoc/tBut	Piperidine	Coupling agent in DMF	TFA	DMF

Common protecting scheme-specific solvents .

The act of removing protecting groups, especially under acidic conditions, results in the production of cationic species that can alkylate the functional groups on the peptide chain. Therefore, scavengers such as water, anisol or thiol derivatives can be added in excess during the deprotection step to react with any of these free reactive species.

Amino Acid Physical Properties

Amino acid coupling

Synthetic peptide coupling requires the activation of the C-terminal carboxylic acid on the incoming amino acid using carbodiimides such as dicyclohexylcarbodiimide (DCC) or diisopropylcarbodiimide (DIC). These coupling reagents react with the carboxyl group to form a highly reactive O-acylisourea intermediate that is quickly displaced by nucleophilic attack from the deprotected primary amino group on the N-terminus of the growing peptide chain to form the nascent peptide bond.

Carbodiimides form such a reactive intermediate that racemization of the amino acid can occur. Therefore, reagents that react with the O-acylisourea intermediate are often added, including 1-hydroxybenzotriazole (HOBt), which forms a less-reactive intermediate that reduces the risk of racemization. Additionally, side reactions caused by carbodiimides have led to the examination of other coupling agents, including benzotriazol-1-yl-oxy-tris(dimethylamino)phosphonium hexafluorophosphate (BOP) and 2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HBTU), which both require activating bases to mediate amino acid coupling.

Peptide cleavage

After successive cycles of amino acid deprotection and coupling, all remaining protecting groups must be removed from the nascent peptide. These groups are cleaved by acidolysis, and the chemical used for cleavage depends on the protection scheme used; strong acids such as hydrogen fluoride (HF), hydrogen bromide (HBr) or trifluoromethane sulfonic acid (TFMSA) are used to cleave Boc and Bzl groups, while a relatively milder acid such as TFA is used to cleave Fmoc and tBut groups. When properly executed, cleavage results in the removal of the N-terminal protecting group of the last amino acid added, the C-terminal protecting group (either chemical or resin) from the first amino acid and any side-chain protecting groups. As with deprotection, scavengers are also included during this step to react with free protecting groups. Because of the importance of cleavage in proper peptide synthesis, this step should be optimized to avoid acid-catalyzed side reactions.

Diagram of peptide cleavage after synthesis. The remaining N-terminal protecting groups, all side-chain protecting groups and the C-terminal protecting group or solid support are removed by strong acid treatment after peptide synthesis is completed.

Liquid-phase peptide synthesis is the classical method that scientists used when first discovering how to generate peptides in vitro and it is still commonly used for large-scale synthesis. This method is slow and labor-intensive, though, because the product has to be manually removed from the reaction solution after each step. Additionally, this approach requires another chemical group to protect the C-terminus of the first amino acid. A benefit of liquid-phase synthesis, though, is that because the product is purified after each step, side reactions are easily detected. Additionally, convergent synthesis can be performed, in which separate peptides are synthesized and then coupled together to create larger peptides.

By far, though, solid-phase peptide synthesis is the most common method of peptide synthesis today. Instead of C-terminal protection with a chemical group, the C-terminus of the first amino acid is coupled to an activated solid support, such as polystyrene or polyacrylamide. This type of approach has a two-fold function: the resin acts as the C-terminal protecting group and provides a rapid method to separate the growing peptide product from the different reaction mixtures during synthesis. As with many different biological manufacturing processes, peptide synthesizers have been developed for automation and high-throughput peptide production.

Although peptide synthesis strategies have been optimized and can be mass-produced, the process to generate peptides is by no means perfect. Events such as incomplete deprotection or reaction with free protecting groups can cause truncated or deletion sequences, isomers or other side products. These events can occur at any step during peptide synthesis, and therefore the longer the peptide sequence, the greater probability that something will negatively affect the synthesis of the target peptide. Thus, peptide yield is inversely correlated with peptide length.

Purification strategies are usually based on a combination of separation methods that exploit the physiochemical characteristics of peptides, including size, charge and hydrophobicity. Purification techniques include:

Size-exclusion chromatography
Ion exchange chromatography (IEC)
Partition chromatography
High-performance liquid chromatography (HPLC)

Reverse-phase chromatography (RPC) is the most versatile and most widely used method of peptide purification. With traditional methods of HPLC, the stationary phase captures polar, hydrophilic molecules that are then differentially eluted by increasing the concentration of polar solvents in the mobile phase. In RPC, as the name implies, hydrophobic molecules from aqueous solutions are instead captured by the stationary phase using hydrophobic C4, C8 or C18 n-alkyl hydrocarbon ligands, and their retention time is a function of the hydrophobicity of the molecule and that of the mobile phase.

For peptide purification, RPC separates the target peptides from impurities from the synthesis steps, such as isomers, deletion sequences, peptide products from side reactions with free coupling and protecting groups or peptides that have undergone side-chain reactions.

Peptide purity is measured as a percentage of the target peptide to impurities that absorb at the peptide bond absorption wavelength (210-220nm), and varying levels of purity are commercially available based on the application in which the peptides will be used:

>95% – Quantitative studies such as NMR, receptor-ligand binding studies, ELISA and RIA, monoclonal antibody production, in vivo studies
>80% – High-throughput screening, non-quantitative blocking in immunohistochemical (IHC) and Western blot analyses, non-quantitative enzyme-substrate studies, antibody affinity purification and plate coating for cell attachment
>70% – ELISA standards, ELISPOT assays and polyclonal antibody production
Protein Isolation and Purification Information
Overview of ELISA
Antibody Production
Overview of Immunohistochemistry (IHC)
Overview of Western Blotting
Enzyme-linked Immunosorbent Assays (ELISA)
Immunohistochemistry
Western Blotting
Lloyd-Williams P. et al. (1997) Chemical approaches to the synthesis of peptides and proteins. Boca Raton: CRC Press. 278
Merrifield R. B. (1963) Solid phase peptide synthesis. I. The synthesis of a tetrapeptide. Journal of the American Chemical Society. 85, 2149-54.
Carpino L. A. (1957) Oxidative reactions of hydrazines. Iv. Elimination of nitrogen from 1, 1-disubstituted-2-arenesulfonhydrazides1-4. Journal of the American Chemical Society. 79, 4427-31.
McKay F. C. and Albertson N. F. (1957) New amine-masking groups for peptide synthesis. Journal of the American Chemical Society. 79, 4686-90.
Anderson G. W. and McGregor A. C. (1957) T-butyloxycarbonylamino acids and their use in peptide synthesis. Journal of the American Chemical Society. 79, 6180-3.
Carpino L. A. and Han G. Y. (1972) 9-fluorenylmethoxycarbonyl amino-protecting group. The Journal of Organic Chemistry. 37, 3404-9.

For Research Use Only. Not for use in diagnostic procedures.

26.7 Peptide Synthesis

26.7 • Peptide Synthesis

Once the structure of a peptide is known, its synthesis can be undertaken—perhaps to obtain a larger amount for biological evaluation. A simple amide might be formed by treating an amine and a carboxylic acid with a carbodiimide (either DCC or EDC; Section 21.3 ), but peptide synthesis is a more difficult problem because many different amide bonds must be formed in a specific order, rather than at random.

The solution to the specificity problem is protection ( Section 17.8 ). If we want to couple alanine with leucine to synthesize Ala-Leu, for instance, we could protect the –NH 2 group of alanine and the –CO 2 H group of leucine to shield them from reacting, then form the desired Ala-Leu amide bond by reaction with EDC or DCC, and then remove the protecting groups.

A number of different amino- and carboxyl-protecting groups have been devised, but only a few are used in peptide synthesis. Carboxyl groups are often protected simply by converting them into methyl or benzyl esters. Both groups are easily introduced by standard methods of ester formation ( Section 21.6 ) and are easily removed by mild hydrolysis with aqueous NaOH. Benzyl esters can also be cleaved by catalytic hydrogenolysis of the weak benzylic C–O bond ( RCO 2 –CH 2 Ph + H 2 → RCO 2 H + PhCH 3 RCO 2 –CH 2 Ph + H 2 → RCO 2 H + PhCH 3 ).

Amino groups are sometimes protected as their tert -butyloxycarbonyl amide (Boc) or, more commonly, as their fluorenylmethyloxycarbonyl amide (Fmoc). The Boc protecting group is introduced by reaction of the amino acid with di- tert -butyl dicarbonate in a nucleophilic acyl substitution reaction and is removed by brief treatment with a strong acid such as trifluoroacetic acid, CF 3 CO 2 H. The Fmoc protecting group is introduced by reaction with fluorenylmethyloxycarbonyl chloride and is removed by treatment with a 20% solution of the amine piperidine in dimethylformamide as solvent.

Thus, five steps are needed to synthesize a dipeptide such as Ala-Leu:

These steps can be repeated to add one amino acid at a time to the growing chain or to link two peptide chains together. Many remarkable achievements in peptide synthesis have been reported, including a complete synthesis of human insulin. Insulin is composed of two chains totaling 51 amino acids linked by two disulfide bridges. The three-dimensional structure of insulin, shown previously, was determined by Dorothy Crowfoot Hodgkin, a British chemist who received the 1964 Nobel Prize in Chemistry for her work on this and other complex biological molecules.

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Home / Introduction to Peptide Synthesis

By James Ashenhurst

Introduction to Peptide Synthesis

Last updated: February 9th, 2023 |

Peptide bonds: Forming peptides from amino acids with the use of protecting groups

Today we’ll go deeper on how to synthesize the most important amides of all – peptides – with an important contribution from protecting group chemistry.

Table of Contents

What Are Peptide Bonds?
The “Proteinogenic” Amino Acids
Synthesis of a Dipeptide Without Protecting Groups
Synthesis of a Dipeptide Using A Protecting Group Strategy
Synthesis of Longer Peptides – Tripeptides and Tetrapeptides
Bonus Topic: Solid-Phase Peptide Synthesis

(Advanced) References and Further Reading

1. what are peptide bonds.

A “peptide bond” is an amide linkage (see Amides: Properties. Synthesis, and Nomenclature ) that connects two amino acids, as in the “dipeptides” L-phenylalanyl-L-valine (below left) and L-leucyl-L-alanine (below right):

2. The “Proteinogenic” Amino Acids

Proteinogenic amino acids are the building blocks of proteins. In addition to the 20 amino acids directly encoded by the genome, two other amino acids are coded into proteins under special circumstances: selenocysteine (present in eukaryotes, including humans) and pyrrolysine (found only in methane-producing bacteria).

With the exception of (achiral) glycine, all proteinogenic amino acids are L-amino acids, where the “L-” prefix relates the stereochemistry of the amino acid relative to that of L-glyceraldehyde [See post: D and L Sugars ] .

Of the chiral amino acids, all are S , with the exception of cysteine and selenocysteine [ Note 1 ] (because sulfur and selenium have a higher priority under the CIP system. )

3. Synthesis of A Simple Dipeptide Without Protecting Groups (is not advisable!)

Let’s build a simple dipeptide between two of these amino acids. For simplicity’s sake, we’ll pick two from the “hydrophobic sidechain” group, alanine (Ala) and leucine (Leu), since their sidechains don’t need additional protecting groups.

What do we need to do to make L-Ala-L-Leu ?

Surveying the methods previously covered to make amides, it might seem simple.

Why not take 1 equivalent each of L-alanine and L-phenylalanine, add a coupling agent like N,N -dicyclohexylcarbodiimide (DCC) and patiently wait for our product to appear?

What could possibly go wrong?

Well, this will give us some of our desired product. But it won’t do so efficiently!

That’s because each amino acid has two reactive termini – an amine and a carboxylic acid – and they can bond together in multiple ways.

Just like the letters A and L can combine to make the words AL and LA, in addition to Ala-Leu (our desired product) we will also get Leu-Ala.

Furthermore, since we’re not adding single molecules together but molar quantities (even a millionth of a mole (a “micromole”) has 10 17 molecules in it) we also have the possibility of forming the “homo-dipeptides” AA (Ala-Ala) and LL (Leu-Leu).

And that’s just a start. No matter how you slice it you’re looking at a low yield (<25%) of the desired material.

That’s inefficient, wasteful, and expensive! Isn’t there a better way?

4. Synthesis Of A Dipeptide Using A Protecting Group Strategy

Yes. Rather than using the native amino acids and just praying for a good yield, we can use protected versions of L-Alanine and L-Leucine.

If we protect the carboxylic acid of Leucine as an ester (e.g. a methyl ester) and protect the amine of L-Alanine as a carbamate (See: Carbamates as protecting groups ) then we set up a situation where we have a single nucleophile and a single electrophile.

This results in a high yield (>95%) of a single product!

[Note: the Boc group is a popular carbamate protecting group for amines; “Boc” stands for t-butyloxycarbonyl]

5. Synthesis of Longer Peptides – Tripeptides and Tetrapeptides

The good news is that we don’t have to stop at the dipeptide. If we choose protecting groups that can be removed selectively (and the carbamate / ester pair qualifies) then we can then deprotect the carbamate, and add a third amino acid.

The choice of carbamate protecting group here was t -butoxycarbonyl (Boc) which is removed with strong acid (trifluoroacetic acid, abbreviated as TFA).

Treatment with TFA removes the Boc group but leaves the methyl ester alone.

So if we treat the dipeptide with TFA, we liberate the amine nitrogen, and can react with another Boc-protected amino acid in the presence of DCC to get a tripeptide.

If we’re keen, we can even extend the same method to build a tetrapeptide, a pentapeptide… or beyond!

It’s not unreasonable to consider this method for longer peptides.

For instance, take something like bradykinin , a 9-peptide chain that causes dilation of blood vessels leading to a rapid drop in blood pressure. (Your body releases bradykinin in response to snake bites, which is how it was originally discovered.)

It might be interesting to synthesize variants of bradykinin where some of the amino acids are swapped out for other ones. In order to do that, we’d need to be able to synthesize it.

So how effective could it be?

If each peptide coupling step has a yield of about 95%, then our overall yield for making bradykinin would be (0.95) 9 , or 63%. That’s actually pretty good! A lot of chemists would be happy to get a yield of 63% for a single reaction, let me tell you.

If the yields are high enough, one can even imagine building something crazy like insulin (51 peptide residues). That’s 7% yield for 51 steps.

Is this possible?

Yes… but it requires a clever modification that won its inventor, Bruce Merrifield, the 1984 Nobel Prize in Chemistry.

6. Bonus Topic: Solid-Phase Peptide Synthesis

What follows below is more supplemental than anything else, but given the importance of the topic, both interesting and useful.

In 1963 a chemist at Rockefeller University named Bruce Merrifield published a paper that would revolutionize how peptides were synthesized, and eventually make the synthesis of long peptides routine.

It was entitled: “ Solid-Phase Peptide Synthesis. I. The Synthesis of a Tetrapeptide “.

Here’s the key idea.

Recall that in our original scheme (above) we protected the carboxylic acid as a methyl ester, which stays the same throughout the whole peptide synthesis.

Merrifield’s idea was: what if we find a way to attach the carboxylic acid to a functional group that is itself linked to a polymer bead ? Not only would this also protect the carboxylic acid, it would drastically improve the ease of separations.

Why? Because instead of having to purify the final product by crystallization or column chromatography, you purify by filtering off the polymer beads (each 200-500 μm) and washing them to remove excess reagents.

The polymer beads themselves are pretty small. A typical size is 200 micrometers. Each bead can load about 4 nanomoles of amino acid.

[ Brandon Finlay from ChemTips shows a picture here ]

The video posted below is not mine, but it gives you an idea of the process.

At about 0:34 you can see how small the beads are.

The starting point for the Merrifield process is crosslinked polystyrene. which behaves like one big interlinked molecule. Polystyrene is then attached to a linker, which usually terminates with an NH 2 group. This itself is usually protected; in order to activate the linker, you need to remove the protecting group cap.

The polymer bead needs to swell in a solvent in order for functional groups on the solid support to undergo reactions efficiently.

The essential procedure is: swell –> add reagents –> wait –> filter –> wash, and repeat. Beads stay in the reaction vessel the whole time. There’s also usually some kind of capping step to make sure any unreacted amines don’t participate in the next reaction.

It’s possible to make peptides up to about 50 units this way. In highly automated systems one can be even more ambitious.

Merrifield started knocking off peptides in the 1960s. Bradykinin was made in 8 days and 68% overall yield. one example. Insulin was made two years later. The crowning achievement of this initial period was probably ribonuclease A , which has 150 amino acid residues.

The original Merrifield process has been significantly modified and improved. Originally, removal of the linker required harsh conditions (strong acid). Today, procedures usually employ FMOC protecting groups instead of BOC, which allow for deprotection with mild amine base (piperidine). A galaxy of new resins, linkers, and coupling procedures have been subsequently developed. The Wikipedia article on solid-phase peptide synthesis is an OK place to start.

Reactions of Diazonium Salts: Sandmeyer and Related Reactions
Protecting Groups for Amines – Carbamates
The Amide Functional Group: Properties, Synthesis, and Nomenclature
Formation of Amides Using DCC (MOC Reaction Guide)
Amine Protection and Deprotection (MOC Reaction Guide)
Amine Practice Questions (MOC Membership)
Isoelectric Points of Amino Acids (and How To Calculate Them)

Note 1 . Cysteine (and selenocysteine) are L, but their stereocenters are (R) , because sulfur and selenium have higher priority within the Cahn-Ingold-Prelog system.

This is a major topic, as the synthesis of peptides is a global billion-dollar industry.

THE SYNTHESIS OF AN OCTAPEPTIDE AMIDE WITH THE HORMONAL ACTIVITY OF OXYTOCIN Vincent du Vigneaud, Charlotte Ressler, College John M. Swan, Carleton W. Roberts, Panayotis G. Katsoyannis, and Samuel Gordon Journal of the American Chemical Society 1953, 75 (19), 4879-4880 DOI: 1021/ja01115a553
The Synthesis of Oxytocin Vincent du Vigneaud, Charlotte Ressler, John M. Swan, Carleton W. Roberts, and Panayotis G. Katsoyannis Journal of the American Chemical Society 1954, 76 (12), 3115-3121 DOI: 1021/ja01641a004
A Method of Synthesis of Long Peptide Chains Using a Synthesis of Oxytocin as an Example Miklos Bodanszky and Vincent du Vigneaud Journal of the American Chemical Society 1959, 81 (21), 5688-5691 DOI: 1021/ja01530a040 In the first half of the 20 th century, peptide synthesis was done using standard organic chemistry solution phase techniques. This is now known as LPPS (liquid-phase peptide synthesis). du Vigneaud received the Nobel Prize in chemistry in 1955 for his work in showing that peptide synthesis could be achieved, using the correct choice of protecting groups and synthetic strategies.In 1963, Prof. Robert Bruce Merrifield (Rockefeller U., New York) revolutionized peptide synthesis by coming up with the SPPS (Solid-Phase Peptide Synthesis) method, making the synthesis of long peptide chains much more feasible. The C-terminus is bound to a polymer resin, and the amino acids are added one at a time following the same cycle: deprotect, wash, couple the next amino acid (with a peptide-coupling reagent such as DCC), wash, deprotect the N-terminus again, and so on. Merrifield’s method came to be called the Boc/Bzl strategy due to the protecting groups employed (Boc for the nitrogen atoms, and Bzl (benzyl) for the side chains). The catch is that final cleavage of the peptide from the resin using this method requires anhydrous HF, which is not easy to handle.
Solid Phase Peptide Synthesis. I. The Synthesis of a Tetrapeptide R. B. Merrifield Journal of the American Chemical Society 1963, 85 (14), 2149-2154 DOI : 10.1021/ja00897a025 This is the paper that started it all – a single-author publication by Prof. Merrifield using the SPPS method to make a tetrapeptide. This remains one of the most highly-viewed and highly-cited papers in JACS , even today.
The Synthesis of Bovine Insulin by the Solid Phase Method Marglin and R. B. Merrifield Journal of the American Chemical Society 1966, 88 (21), 5051-5052 DOI: 10.1021/ja00973a068 Insulin is a billion-dollar hormone as its dysregulation is what causes diabetes. This paper shows that insulin can be made through SPPS methods. Insulin is tricky to make as it has 2 chains (the A and B chain) connected through disulfide bonds. Interestingly, Merrifield synthesized the active hormone by combining both chains with protected thiols (protected as sulfonates), which he then reduced to the thiol, and then oxidized in air in a basic medium (pH 10). This is called undirected or air oxidation since the disulfide bonds are not being formed selectively; luckily they formed correctly in this case. Insulin today is not manufactured by the SPPS method due to these complications; instead it is made through a recombinant process.
The Synthesis of Ribonuclease A Bernd Gutte and R. B. Merrifield Journal of Biological Chemistry 1971 , 246 (6), 1922-1941 DOI: 10.1016/S0021-9258(18)62396-8 This is the crowning achievement of SPPS – the synthesis of a 124-mer peptide (or protein, at this point), RNAse A.
The Solid Phase Synthesis of Ribonuclease A by Robert Bruce Merrifield Nicole Kresge, Robert D. Simoni and Robert L. Hill Journal of Biological Chemistry 2006 , 281 (26), e21 DOI: 10.1016/S0021-9258(20)55702-5 A short biographical account of Prof. Merrifield’s life. This mentions that he developed the first prototype of an automated peptide synthesizer working in the basement of his house in 1965!
Solid Phase Synthesis Nobel Lecture , December 8, 1984 by Bruce Merrifield Merrifield’s Nobel Lecture upon receiving the Nobel Prize in Chemistry in 1984. This describes his life, how he conceived of and developed the SPPS process, and all the breakthroughs it has enabled.
9-Fluorenylmethoxycarbonyl function, a new base-sensitive amino-protecting group Louis A. Carpino and Grace Y. Han Journal of the American Chemical Society 1970, 92 (19), 5748-5749 DOI : 10.1021/ja00722a043
9-Fluorenylmethoxycarbonyl amino-protecting group Louis A. Carpino and Grace Y. Han The Journal of Organic Chemistry 1972 37 (22), 3404-3409 DOI : 10.1021/jo00795a005 The discovery and development of the Fmoc (9-fluorenylmethoxycarbonyl) group as a protecting group for amines has also improved the practice of peptide synthesis and SPPS.
A mild procedure for solid phase peptide synthesis: use of fluorenylmethoxycarbonylamino-acids Atherton, Hazel Fox, Diana Harkiss, C. J. Logan, R. C. Sheppard and B. J. Williams J. Chem. Soc. Chem. Comm. 1978 , 537-539 DOI : 10.1039/C39780000537 This is the first paper describing what is now known as the Fmoc/tBu SPPS process, which has largely supplanted the original Boc/Bzl process developed by Prof. Merrifield. The advantages with Fmoc-SPPS are multifold, but the main ones are simplicity and orthogonality. Amine deprotection is done with a base (20% piperidine in DMF is sufficient to deprotect an Fmoc-amine), and final cleavage of the peptide from the resin can be done with TFA (trifluoroacetic acid), which is much easier to handle than HF.
Advances in Fmoc solid-phase peptide synthesis Raymond Behrendt, Peter White, and John Offer Peptide Sci. 2016, 22 , 4-27 DOI: 10.1002/psc.2836 A modern review on Fmoc-SPPS that describes how far we have come and some of the challenges that remain. For instance, aggregation of peptide chains on the resin is a major issue in Fmoc-SPPS when synthesizing especially hydrophobic peptides, and there are ways to deal with it, such as the introduction of ‘kinking’ residues, like Pro or the use of pseudoprolines, which will revert back to the desired amino acids when the peptide is cleaved with TFA. Native Chemical Ligation is also used nowadays for the synthesis of especially long peptides, like Merrifield’s RNAse A.

00 General Chemistry Review

Lewis Structures
Ionic and Covalent Bonding
Chemical Kinetics
Chemical Equilibria
Valence Electrons of the First Row Elements
How Concepts Build Up In Org 1 ("The Pyramid")

01 Bonding, Structure, and Resonance

How Do We Know Methane (CH4) Is Tetrahedral?
Hybrid Orbitals and Hybridization
How To Determine Hybridization: A Shortcut
Orbital Hybridization And Bond Strengths
Sigma bonds come in six varieties: Pi bonds come in one
A Key Skill: How to Calculate Formal Charge
The Four Intermolecular Forces and How They Affect Boiling Points
3 Trends That Affect Boiling Points
How To Use Electronegativity To Determine Electron Density (and why NOT to trust formal charge)
Introduction to Resonance
How To Use Curved Arrows To Interchange Resonance Forms
Evaluating Resonance Forms (1) - The Rule of Least Charges
How To Find The Best Resonance Structure By Applying Electronegativity
Evaluating Resonance Structures With Negative Charges
Evaluating Resonance Structures With Positive Charge
Exploring Resonance: Pi-Donation
Exploring Resonance: Pi-acceptors
In Summary: Evaluating Resonance Structures
Drawing Resonance Structures: 3 Common Mistakes To Avoid
How to apply electronegativity and resonance to understand reactivity
Bond Hybridization Practice
Structure and Bonding Practice Quizzes
Resonance Structures Practice

02 Acid Base Reactions

Introduction to Acid-Base Reactions
Acid Base Reactions In Organic Chemistry
The Stronger The Acid, The Weaker The Conjugate Base
Walkthrough of Acid-Base Reactions (3) - Acidity Trends
Five Key Factors That Influence Acidity
Acid-Base Reactions: Introducing Ka and pKa
How to Use a pKa Table
The pKa Table Is Your Friend
A Handy Rule of Thumb for Acid-Base Reactions
Acid Base Reactions Are Fast
pKa Values Span 60 Orders Of Magnitude
How Protonation and Deprotonation Affect Reactivity
Acid Base Practice Problems

03 Alkanes and Nomenclature

Meet the (Most Important) Functional Groups
Condensed Formulas: Deciphering What the Brackets Mean
Hidden Hydrogens, Hidden Lone Pairs, Hidden Counterions
Don't Be Futyl, Learn The Butyls
Primary, Secondary, Tertiary, Quaternary In Organic Chemistry
Branching, and Its Affect On Melting and Boiling Points
The Many, Many Ways of Drawing Butane
Wedge And Dash Convention For Tetrahedral Carbon
Common Mistakes in Organic Chemistry: Pentavalent Carbon
Table of Functional Group Priorities for Nomenclature
Summary Sheet - Alkane Nomenclature
Organic Chemistry IUPAC Nomenclature Demystified With A Simple Puzzle Piece Approach
Boiling Point Quizzes
Organic Chemistry Nomenclature Quizzes

04 Conformations and Cycloalkanes

Staggered vs Eclipsed Conformations of Ethane
Conformational Isomers of Propane
Newman Projection of Butane (and Gauche Conformation)
Introduction to Cycloalkanes (1)
Geometric Isomers In Small Rings: Cis And Trans Cycloalkanes
Calculation of Ring Strain In Cycloalkanes
Cycloalkanes - Ring Strain In Cyclopropane And Cyclobutane
Cyclohexane Conformations
Cyclohexane Chair Conformation: An Aerial Tour
How To Draw The Cyclohexane Chair Conformation
The Cyclohexane Chair Flip
The Cyclohexane Chair Flip - Energy Diagram
Substituted Cyclohexanes - Axial vs Equatorial
Ranking The Bulkiness Of Substituents On Cyclohexanes: "A-Values"
Cyclohexane Chair Conformation Stability: Which One Is Lower Energy?
Fused Rings - Cis-Decalin and Trans-Decalin
Naming Bicyclic Compounds - Fused, Bridged, and Spiro
Bredt's Rule (And Summary of Cycloalkanes)
Newman Projection Practice
Cycloalkanes Practice Problems

05 A Primer On Organic Reactions

The Most Important Question To Ask When Learning a New Reaction
Learning New Reactions: How Do The Electrons Move?
The Third Most Important Question to Ask When Learning A New Reaction
7 Factors that stabilize negative charge in organic chemistry
7 Factors That Stabilize Positive Charge in Organic Chemistry
Nucleophiles and Electrophiles
Curved Arrows (for reactions)
Curved Arrows (2): Initial Tails and Final Heads
Nucleophilicity vs. Basicity
The Three Classes of Nucleophiles
What Makes A Good Nucleophile?
What makes a good leaving group?
3 Factors That Stabilize Carbocations
Equilibrium and Energy Relationships
What's a Transition State?
Hammond's Postulate
Learning Organic Chemistry Reactions: A Checklist (PDF)
Introduction to Free Radical Substitution Reactions
Introduction to Oxidative Cleavage Reactions

06 Free Radical Reactions

Bond Dissociation Energies = Homolytic Cleavage
Free Radical Reactions
3 Factors That Stabilize Free Radicals
What Factors Destabilize Free Radicals?
Bond Strengths And Radical Stability
Free Radical Initiation: Why Is "Light" Or "Heat" Required?
Initiation, Propagation, Termination
Monochlorination Products Of Propane, Pentane, And Other Alkanes
Selectivity In Free Radical Reactions
Selectivity in Free Radical Reactions: Bromination vs. Chlorination
Halogenation At Tiffany's
Allylic Bromination
Bonus Topic: Allylic Rearrangements
In Summary: Free Radicals
Synthesis (2) - Reactions of Alkanes
Free Radicals Practice Quizzes

07 Stereochemistry and Chirality

Types of Isomers: Constitutional Isomers, Stereoisomers, Enantiomers, and Diastereomers
How To Draw The Enantiomer Of A Chiral Molecule
How To Draw A Bond Rotation
Introduction to Assigning (R) and (S): The Cahn-Ingold-Prelog Rules
Assigning Cahn-Ingold-Prelog (CIP) Priorities (2) - The Method of Dots
Enantiomers vs Diastereomers vs The Same? Two Methods For Solving Problems
Assigning R/S To Newman Projections (And Converting Newman To Line Diagrams)
How To Determine R and S Configurations On A Fischer Projection
The Meso Trap
Optical Rotation, Optical Activity, and Specific Rotation
Optical Purity and Enantiomeric Excess
What's a Racemic Mixture?
Chiral Allenes And Chiral Axes
Stereochemistry Practice Problems and Quizzes

08 Substitution Reactions

Introduction to Nucleophilic Substitution Reactions
Walkthrough of Substitution Reactions (1) - Introduction
Two Types of Nucleophilic Substitution Reactions
The SN2 Mechanism
Why the SN2 Reaction Is Powerful
The SN1 Mechanism
The Conjugate Acid Is A Better Leaving Group
Comparing the SN1 and SN2 Reactions
Polar Protic? Polar Aprotic? Nonpolar? All About Solvents
Steric Hindrance is Like a Fat Goalie
Common Blind Spot: Intramolecular Reactions
The Conjugate Base is Always a Stronger Nucleophile
Substitution Practice - SN1
Substitution Practice - SN2

09 Elimination Reactions

Elimination Reactions (1): Introduction And The Key Pattern
Elimination Reactions (2): The Zaitsev Rule
Elimination Reactions Are Favored By Heat
Two Elimination Reaction Patterns
The E1 Reaction
The E2 Mechanism
E1 vs E2: Comparing the E1 and E2 Reactions
Antiperiplanar Relationships: The E2 Reaction and Cyclohexane Rings
Bulky Bases in Elimination Reactions
Comparing the E1 vs SN1 Reactions
Elimination (E1) Reactions With Rearrangements
E1cB - Elimination (Unimolecular) Conjugate Base
Elimination (E1) Practice Problems And Solutions
Elimination (E2) Practice Problems and Solutions

10 Rearrangements

Introduction to Rearrangement Reactions
Rearrangement Reactions (1) - Hydride Shifts
Carbocation Rearrangement Reactions (2) - Alkyl Shifts
Pinacol Rearrangement
The SN1, E1, and Alkene Addition Reactions All Pass Through A Carbocation Intermediate

11 SN1/SN2/E1/E2 Decision

Identifying Where Substitution and Elimination Reactions Happen
Deciding SN1/SN2/E1/E2 (1) - The Substrate
Deciding SN1/SN2/E1/E2 (2) - The Nucleophile/Base
SN1 vs E1 and SN2 vs E2 : The Temperature
Deciding SN1/SN2/E1/E2 - The Solvent
Wrapup: The Key Factors For Determining SN1/SN2/E1/E2
Alkyl Halide Reaction Map And Summary
SN1 SN2 E1 E2 Practice Problems

12 Alkene Reactions

E and Z Notation For Alkenes (+ Cis/Trans)
Alkene Stability
Alkene Addition Reactions: "Regioselectivity" and "Stereoselectivity" (Syn/Anti)
Stereoselective and Stereospecific Reactions
Hydrohalogenation of Alkenes and Markovnikov's Rule
Hydration of Alkenes With Aqueous Acid
Rearrangements in Alkene Addition Reactions
Halogenation of Alkenes and Halohydrin Formation
Oxymercuration Demercuration of Alkenes
Hydroboration Oxidation of Alkenes
m-CPBA (meta-chloroperoxybenzoic acid)
OsO4 (Osmium Tetroxide) for Dihydroxylation of Alkenes
Palladium on Carbon (Pd/C) for Catalytic Hydrogenation of Alkenes
Cyclopropanation of Alkenes
A Fourth Alkene Addition Pattern - Free Radical Addition
Alkene Reactions: Ozonolysis
Summary: Three Key Families Of Alkene Reaction Mechanisms
Synthesis (4) - Alkene Reaction Map, Including Alkyl Halide Reactions
Alkene Reactions Practice Problems

13 Alkyne Reactions

Acetylides from Alkynes, And Substitution Reactions of Acetylides
Partial Reduction of Alkynes With Lindlar's Catalyst
Partial Reduction of Alkynes With Na/NH3 To Obtain Trans Alkenes
Alkyne Hydroboration With "R2BH"
Hydration and Oxymercuration of Alkynes
Hydrohalogenation of Alkynes
Alkyne Halogenation: Bromination, Chlorination, and Iodination of Alkynes
Alkyne Reactions - The "Concerted" Pathway
Alkenes To Alkynes Via Halogenation And Elimination Reactions
Alkynes Are A Blank Canvas
Synthesis (5) - Reactions of Alkynes
Alkyne Reactions Practice Problems With Answers

14 Alcohols, Epoxides and Ethers

Alcohols - Nomenclature and Properties
Alcohols Can Act As Acids Or Bases (And Why It Matters)
Alcohols - Acidity and Basicity
The Williamson Ether Synthesis
Ethers From Alkenes, Tertiary Alkyl Halides and Alkoxymercuration
Alcohols To Ethers via Acid Catalysis
Cleavage Of Ethers With Acid
Epoxides - The Outlier Of The Ether Family
Opening of Epoxides With Acid
Epoxide Ring Opening With Base
Making Alkyl Halides From Alcohols
Tosylates And Mesylates
PBr3 and SOCl2
Elimination Reactions of Alcohols
Elimination of Alcohols To Alkenes With POCl3
Alcohol Oxidation: "Strong" and "Weak" Oxidants
Demystifying The Mechanisms of Alcohol Oxidations
Protecting Groups For Alcohols
Thiols And Thioethers
Calculating the oxidation state of a carbon
Oxidation and Reduction in Organic Chemistry
Oxidation Ladders
SOCl2 Mechanism For Alcohols To Alkyl Halides: SN2 versus SNi
Alcohol Reactions Roadmap (PDF)
Alcohol Reaction Practice Problems
Epoxide Reaction Quizzes
Oxidation and Reduction Practice Quizzes

15 Organometallics

What's An Organometallic?
Formation of Grignard and Organolithium Reagents
Organometallics Are Strong Bases
Reactions of Grignard Reagents
Protecting Groups In Grignard Reactions
Synthesis Problems Involving Grignard Reagents
Grignard Reactions And Synthesis (2)
Organocuprates (Gilman Reagents): How They're Made
Gilman Reagents (Organocuprates): What They're Used For
The Heck, Suzuki, and Olefin Metathesis Reactions (And Why They Don't Belong In Most Introductory Organic Chemistry Courses)
Reaction Map: Reactions of Organometallics
Grignard Practice Problems

16 Spectroscopy

Degrees of Unsaturation (or IHD, Index of Hydrogen Deficiency)
Conjugation And Color (+ How Bleach Works)
Introduction To UV-Vis Spectroscopy
UV-Vis Spectroscopy: Absorbance of Carbonyls
UV-Vis Spectroscopy: Practice Questions
Bond Vibrations, Infrared Spectroscopy, and the "Ball and Spring" Model
Infrared Spectroscopy: A Quick Primer On Interpreting Spectra
IR Spectroscopy: 4 Practice Problems
1H NMR: How Many Signals?
Homotopic, Enantiotopic, Diastereotopic
Diastereotopic Protons in 1H NMR Spectroscopy: Examples
C13 NMR - How Many Signals
Liquid Gold: Pheromones In Doe Urine
Natural Product Isolation (1) - Extraction
Natural Product Isolation (2) - Purification Techniques, An Overview
Structure Determination Case Study: Deer Tarsal Gland Pheromone

17 Dienes and MO Theory

What To Expect In Organic Chemistry 2
Are these molecules conjugated?
Conjugation And Resonance In Organic Chemistry
Bonding And Antibonding Pi Orbitals
Molecular Orbitals of The Allyl Cation, Allyl Radical, and Allyl Anion
Pi Molecular Orbitals of Butadiene
Reactions of Dienes: 1,2 and 1,4 Addition
Thermodynamic and Kinetic Products
More On 1,2 and 1,4 Additions To Dienes
s-cis and s-trans
The Diels-Alder Reaction
Cyclic Dienes and Dienophiles in the Diels-Alder Reaction
Stereochemistry of the Diels-Alder Reaction
Exo vs Endo Products In The Diels Alder: How To Tell Them Apart
HOMO and LUMO In the Diels Alder Reaction
Why Are Endo vs Exo Products Favored in the Diels-Alder Reaction?
Diels-Alder Reaction: Kinetic and Thermodynamic Control
The Retro Diels-Alder Reaction
The Intramolecular Diels Alder Reaction
Regiochemistry In The Diels-Alder Reaction
The Cope and Claisen Rearrangements
Electrocyclic Reactions
Electrocyclic Ring Opening And Closure (2) - Six (or Eight) Pi Electrons
Diels Alder Practice Problems
Molecular Orbital Theory Practice

18 Aromaticity

Introduction To Aromaticity
Rules For Aromaticity
Huckel's Rule: What Does 4n+2 Mean?
Aromatic, Non-Aromatic, or Antiaromatic? Some Practice Problems
Antiaromatic Compounds and Antiaromaticity
The Pi Molecular Orbitals of Benzene
The Pi Molecular Orbitals of Cyclobutadiene
Frost Circles
Aromaticity Practice Quizzes

19 Reactions of Aromatic Molecules

Electrophilic Aromatic Substitution: Introduction
Activating and Deactivating Groups In Electrophilic Aromatic Substitution
Electrophilic Aromatic Substitution - The Mechanism
Ortho-, Para- and Meta- Directors in Electrophilic Aromatic Substitution
Understanding Ortho, Para, and Meta Directors
Why are halogens ortho- para- directors?
Disubstituted Benzenes: The Strongest Electron-Donor "Wins"
Electrophilic Aromatic Substitutions (1) - Halogenation of Benzene
Electrophilic Aromatic Substitutions (2) - Nitration and Sulfonation
EAS Reactions (3) - Friedel-Crafts Acylation and Friedel-Crafts Alkylation
Intramolecular Friedel-Crafts Reactions
Nucleophilic Aromatic Substitution (NAS)
Nucleophilic Aromatic Substitution (2) - The Benzyne Mechanism
Reactions on the "Benzylic" Carbon: Bromination And Oxidation
The Wolff-Kishner, Clemmensen, And Other Carbonyl Reductions
More Reactions on the Aromatic Sidechain: Reduction of Nitro Groups and the Baeyer Villiger
Aromatic Synthesis (1) - "Order Of Operations"
Synthesis of Benzene Derivatives (2) - Polarity Reversal
Aromatic Synthesis (3) - Sulfonyl Blocking Groups
Birch Reduction
Synthesis (7): Reaction Map of Benzene and Related Aromatic Compounds
Aromatic Reactions and Synthesis Practice
Electrophilic Aromatic Substitution Practice Problems

20 Aldehydes and Ketones

What's The Alpha Carbon In Carbonyl Compounds?
Nucleophilic Addition To Carbonyls
Aldehydes and Ketones: 14 Reactions With The Same Mechanism
Sodium Borohydride (NaBH4) Reduction of Aldehydes and Ketones
Grignard Reagents For Addition To Aldehydes and Ketones
Wittig Reaction
Hydrates, Hemiacetals, and Acetals
Imines - Properties, Formation, Reactions, and Mechanisms
All About Enamines
Breaking Down Carbonyl Reaction Mechanisms: Reactions of Anionic Nucleophiles (Part 2)
Aldehydes Ketones Reaction Practice

21 Carboxylic Acid Derivatives

Nucleophilic Acyl Substitution (With Negatively Charged Nucleophiles)
Addition-Elimination Mechanisms With Neutral Nucleophiles (Including Acid Catalysis)
Basic Hydrolysis of Esters - Saponification
Transesterification
Proton Transfer
Fischer Esterification - Carboxylic Acid to Ester Under Acidic Conditions
Lithium Aluminum Hydride (LiAlH4) For Reduction of Carboxylic Acid Derivatives
LiAlH[Ot-Bu]3 For The Reduction of Acid Halides To Aldehydes
Di-isobutyl Aluminum Hydride (DIBAL) For The Partial Reduction of Esters and Nitriles
Amide Hydrolysis
Thionyl Chloride (SOCl2)
Diazomethane (CH2N2)
Carbonyl Chemistry: Learn Six Mechanisms For the Price Of One
Making Music With Mechanisms (PADPED)
Carboxylic Acid Derivatives Practice Questions

22 Enols and Enolates

Keto-Enol Tautomerism
Enolates - Formation, Stability, and Simple Reactions
Kinetic Versus Thermodynamic Enolates
Aldol Addition and Condensation Reactions
Reactions of Enols - Acid-Catalyzed Aldol, Halogenation, and Mannich Reactions
Claisen Condensation and Dieckmann Condensation
Decarboxylation
The Malonic Ester and Acetoacetic Ester Synthesis
The Michael Addition Reaction and Conjugate Addition
The Robinson Annulation
Haloform Reaction
The Hell–Volhard–Zelinsky Reaction
Enols and Enolates Practice Quizzes
Basicity of Amines And pKaH
5 Key Basicity Trends of Amines
The Mesomeric Effect And Aromatic Amines
Nucleophilicity of Amines
Alkylation of Amines (Sucks!)
Reductive Amination
The Gabriel Synthesis
Some Reactions of Azides
The Hofmann Elimination
The Hofmann and Curtius Rearrangements
The Cope Elimination
Protecting Groups for Amines - Carbamates
The Strecker Synthesis of Amino Acids
Amine Practice Questions

24 Carbohydrates

D and L Notation For Sugars
Pyranoses and Furanoses: Ring-Chain Tautomerism In Sugars
What is Mutarotation?
Reducing Sugars
The Big Damn Post Of Carbohydrate-Related Chemistry Definitions
The Haworth Projection
Converting a Fischer Projection To A Haworth (And Vice Versa)
Reactions of Sugars: Glycosylation and Protection
The Ruff Degradation and Kiliani-Fischer Synthesis
Carbohydrates Practice
Amino Acid Quizzes

25 Fun and Miscellaneous

A Gallery of Some Interesting Molecules From Nature
Screw Organic Chemistry, I'm Just Going To Write About Cats
On Cats, Part 1: Conformations and Configurations
On Cats, Part 2: Cat Line Diagrams
On Cats, Part 4: Enantiocats
On Cats, Part 6: Stereocenters
Organic Chemistry Is Shit
The Organic Chemistry Behind "The Pill"
Maybe they should call them, "Formal Wins" ?
Why Do Organic Chemists Use Kilocalories?
The Principle of Least Effort
Organic Chemistry GIFS - Resonance Forms
Reproducibility In Organic Chemistry
What Holds The Nucleus Together?
How Reactions Are Like Music
Organic Chemistry and the New MCAT

26 Organic Chemistry Tips and Tricks

Common Mistakes: Formal Charges Can Mislead
Partial Charges Give Clues About Electron Flow
Draw The Ugly Version First
Organic Chemistry Study Tips: Learn the Trends
The 8 Types of Arrows In Organic Chemistry, Explained
Top 10 Skills To Master Before An Organic Chemistry 2 Final
Common Mistakes with Carbonyls: Carboxylic Acids... Are Acids!
Planning Organic Synthesis With "Reaction Maps"
Alkene Addition Pattern #1: The "Carbocation Pathway"
Alkene Addition Pattern #2: The "Three-Membered Ring" Pathway
Alkene Addition Pattern #3: The "Concerted" Pathway
Number Your Carbons!
The 4 Major Classes of Reactions in Org 1
How (and why) electrons flow
Grossman's Rule
Three Exam Tips
A 3-Step Method For Thinking Through Synthesis Problems
Putting It Together
Putting Diels-Alder Products in Perspective
The Ups and Downs of Cyclohexanes
The Most Annoying Exceptions in Org 1 (Part 1)
The Most Annoying Exceptions in Org 1 (Part 2)
The Marriage May Be Bad, But the Divorce Still Costs Money
9 Nomenclature Conventions To Know
Nucleophile attacks Electrophile

27 Case Studies of Successful O-Chem Students

Success Stories: How Corina Got The The "Hard" Professor - And Got An A+ Anyway
How Helena Aced Organic Chemistry
From a "Drop" To B+ in Org 2 – How A Hard Working Student Turned It Around
How Serge Aced Organic Chemistry
Success Stories: How Zach Aced Organic Chemistry 1
Success Stories: How Kari Went From C– to B+
How Esther Bounced Back From a "C" To Get A's In Organic Chemistry 1 And 2
How Tyrell Got The Highest Grade In Her Organic Chemistry Course
This Is Why Students Use Flashcards
Success Stories: How Stu Aced Organic Chemistry
How John Pulled Up His Organic Chemistry Exam Grades
Success Stories: How Nathan Aced Organic Chemistry (Without It Taking Over His Life)
How Chris Aced Org 1 and Org 2
Interview: How Jay Got an A+ In Organic Chemistry
How to Do Well in Organic Chemistry: One Student's Advice
"America's Top TA" Shares His Secrets For Teaching O-Chem
"Organic Chemistry Is Like..." - A Few Metaphors
How To Do Well In Organic Chemistry: Advice From A Tutor
Guest post: "I went from being afraid of tests to actually looking forward to them".

Comment section

6 thoughts on “ introduction to peptide synthesis ”.

Thanks for very good explanation and excellent step-by-step video!!

These tips are awesome, I was looking for something like that. Thanks!

OK. Do you guys actually do peptide synthesis or do you just drop ship?

We, or least, in my school do cover this topic extensively. We need to learn fischer esterfication of them, acylation, protecting group although it was different than the protecting group you used It had two benzene rings and a five member ring in the middle.

Good to know! Sounds like you’re talking about FMOC. https://en.wikipedia.org/wiki/Fluorenylmethyloxycarbonyl_protecting_group

I should update to include that.

Thank you very much!

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StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2024 Jan-.

StatPearls [Internet].

Biochemistry, peptide.

Jessica Forbes ; Karthik Krishnamurthy .

Affiliations

Last Update: August 28, 2023 .

Introduction

Peptides play an essential role in fundamental physiological processes and are necessary for many biochemical processes. A peptide is a short string of 2 to 50 amino acids, formed by a condensation reaction, joining together through a covalent bond. [1] Sequential covalent bonds with additional amino acids yield a peptide chain and the building block of proteins.

Peptides are named based on the number of amino acid residues in the sequence. As peptide chains form between joining of the primary structure of amino acids, they may enlarge to become an oligopeptide when there are between 10 to 20 amino acids in the chain. In vivo, each amino acid is added to the amino-terminal of one amino acid to form a peptide chain. [1] When there are greater than 20 amino acids, the peptide is an unbranched chain deemed a polypeptide.

Each amino acid comprising a peptide is called a “residue” since that is the portion remaining after the loss of water in the dehydration reaction. Amino acids are the organic starting molecule composed of a carboxyl-terminal and an amino group that makes up the foundation of a protein. Peptide synthesis depends on three main reactions: 1. an amino acid goes through a deprotection step, a preparatory reaction that adds the next amino acid to the chain, and lastly, a coupling reaction that forms the final peptide with functionality. [1] In the second step, the amino acid becomes activated with several reagents. Thes carboxylic acid in the amino acid will react to make the activated form, which will then enter into a coupling reaction. After one round of peptide synthesis, this process is repeatable to add more amino acids until creating the desired length of the peptide.

Peptide bonds are resistant to conditions that denature proteins, such as elevated temperatures and high concentration of urea. Amino acids all have the same general structure, with a positive charge on nitrogen and negative on the carbonyl group. [1]

Fundamentals

Peptide bonds

The peptide bond formed in the active site of the ribosome has a partial double-bond character. [2] This bond is more rigid and planar than a single bond since double bonds are shorter and stronger and require more free energy to break them. Due to the steric interference of R groups, the bond is almost always a trans bond. [2] The nature of the bond prevents complete free rotation between the carbonyl carbon and the nitrogen of the peptide bond. However, the bonds between the other carbon atoms can freely rotate. This configuration and allows for multiple configurations and isomers of peptides to be created.

Bioactive Peptides

As amino acids combine to form a peptide, specific bioactive peptides can be designed with implications to the pharmaceutical industry and biologics design usage for therapeutic biomedical research. [3] Extensive research has shown the multi-faceted role of bioactive peptides has demonstrated effectiveness in blood pressure decreasing, anti-microbial properties, anti-inflammatory, anti-thrombotic, improved response to infection, and anti-oxidant. [4] The fundamental nature of peptides as the building blocks of proteins, allow for the synthetic and in vitro mimicking of these endogenous substances to that regulate specific cellular functions and facilitate an innumerable amount of biochemical process in the body.

Cellular Level

The process of biochemical synthesis of a peptide from its primary amino acid primary structure to a final protein structure is a fundamental biological process. This section will provide an overview of the mechanisms involved in synthesizing a peptide sequence and highlight key cellular locations and specific enzymes.

Biologically active peptides, including neurotransmitters and hormones and, are created from and RNA template, transcribed from DNA. [1] First, a ribosome translates a signal sequence that docks it to a signal recognition particle (SRP) on the rough endoplasmic reticulum (RER). In vivo, after transiting from the nucleus to the cytoplasm to the attachment of a ribosome, mRNA will begin the process of translation and peptide chain formation.

The steps of translation subdivide into initiation, elongation, and termination. [1] The initiation step includes an mRNA binding to a small ribosome subunit. [2] A group of similar nucleotide sequences, termed Kozak sequences surround the start codon; they act as a landmark for small ribosomal subunit to recognize and attach the start codon, AUG, coding for Methionine, binds to the anticodon of tRNA. The large subunit holds an A, P, and E site, and the first step is the binding of the small subunit in the P site. [5] After each amino acid attaches to its corresponding tRNA with the help of ATP, the enzyme aminoacyl-tRNA synthetase catalyzes the bond. [5] As each peptide bond forms, linking together two amino acids, a condensation reaction occurs with the loss of a water molecule. [1]

With expansion and addition of additional amino acids, a polypeptide is then created and destined to become the major macromolecule constituent of cells, protein. Post-translationally modification of peptides such as methylation, phosphorylation, acetylation can also alter the rates of peptide synthesis. [6]

As a growing peptide forms, it is then cleaved from its signal sequence, forming a large preprohormone, which is then cleaved further into a prohormone. As a prohormone, it is packaged into vesicles and sent to the Golgi apparatus for further processing and to be proteolytically cleaved into their final form. The final peptide is packaged into secretory vesicles and sent into the cytoplasm and then leave the cell via exocytosis when they receive a stimulus.

The established method in a laboratory setting for the production of the synthetic peptide is known as solid-phase peptide synthesis (SPPS). [7] This process allows the rapid assembly of a peptide chain through a process of consecutive reactions of amino acid derivatives in a series of coupling, deprotecting techniques. [7] [8]

Molecular Level

Peptide Hormones

Peptide hormones are water-soluble molecules that can range from 3 to 200 amino acids in lengths and shape and are linked by peptide bonds. Peptide hormones are synthesized locally and can travel to remote tissues with an implication for physiological growth and differentiation. The paracrine and perhaps autocrine actions of these peptide hormones contribute to the growth, survival, and functionality of the tissues on which they act. [9] These hormones range broadly in size, structure, and function. The following is only a concise list and does not represent all of the physiologic and endogenous peptide hormones in the body; however, these peptide families are of note.

Pro-opiomelanocortin (POMC) gene family is originally a 241 amino acid residue that is cleaved at different lysine residues through proteolysis to create unique, active peptides. The peptides created include melanocyte-stimulating factor (MSH), adrenocorticotropic releasing hormone (ACTH), B-lipotropin, and B-endorphin, and are expressed in peripheral tissues and the brain. [10]

Oxytocin and ADH The posterior pituitary produces two peptide hormones that differ by only two AAs: oxytocin and anti-diuretic hormone (ADH). Both oxytocin and ADH are nonapeptides with a disulfide bridge. [11] These nonapeptides are packaged through a process involving carrier proteins called neurophysins. [12]

Insulin is a 51 amino acid peptide hormone that consists of two disulfide-linked peptide chains. [12] IGF-1 (insulin-like-growth-factor- 1) family are also peptide hormones but have three disulfide bonds. [12] Insulin's role in the body is multifaceted to control metabolic homeostasis, including glucose uptake from the blood and storage of glucose as glycogen in the liver.

Glucagon is created when proglucagon is cleaved by prohormone convertase 2, to form a fully processed bioactive peptide. [13] It is released by pancreatic alpha cells in response to hypoglycemia or even during a homeostatic increase in concentrations of amino acids. [13] Glucagon's effects to promote homeostatic equilibrium throughout the body, work through mechanisms that balance energy expenditure and glucose metabolism. The study of this peptide hormone and its mechanisms provides a basis of understanding of therapeutics for diabetes management and other conditions. [13]

Secretin is another example of a peptide hormone, with an N-terminal and C-terminal end, composed of a 27 chain amino acid sequence. [14] This peptide originates from the SCT gene, and first becomes a prohormone known as prosecretin. Once activated by exposure to gastric acid, it is cleaved into the active peptide form and released by S cells in the mucosa of the duodenum. [14] It functions to stimulate the pancreas and bile ducts to release bicarbonate, which acts to neutralize potentially harmful gastric acids entering into the stomach. [15]

Calcitonin gene-related peptide (CGRP) is a 37-amino acid neuropeptide, which is most commonly localized to C and Aδ sensory fibers but affects both the central and peripheral nervous systems and metabolism through a multitude of receptor types. [16] New research has demonstrated that intraperitoneal treatment with CGRP can have an energy stimulating effect and even promote an increased appetite. [17] Current research has also linked the activity CGRP in the cerebrovascular system to a possible etiology of migraine attacks. [18] CGRP is mainly found in the enteric nervous system but has been postulated to play a potential role in cranial nociception and cerebral vasodilation, leading to severe migraine headaches. [19]

Natriuretic Peptides are small peptide hormones secreted by cardiac myocytes in response to tension or wall stress. [20] This peptide system, including atrial natriuretic peptides (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP), are all secreted by the cardiac atrium as protective mechanisms to prevent adverse cardiovascular/renal conditions including anti-proliferative, anti-remodeling, vasodilative, and modulation of the renin-angiotensin-aldosterone system. [20] The peptide, ANP, is secreted in the atrium of cardiac tissue and is known to be 28 amino acids in length linked by disulfide bonds. [20] BNP, on the other hand, shares 17 common amino acids with ANP but is a 32 amino acid peptide, while CNP is a total of 22 amino acids in length. [20] Isolation and understanding of these peptide hormones have provided a better understanding of physiologic blood pressure control and an opportunity for natriuretic peptides for therapeutic purposes. [21]

Pathophysiology

Pathophysiological processes related to peptides are very broad due to the ubiquitous nature of peptides in the body. This section will describe how peptides are involved in the pathophysiology of various metabolic processes.

Peptide-Receptor Complex and Signaling Cascade

Biologically active peptides are produced from genes that target specific proteins or protein-coupled receptors, such as G-protein-coupled-receptors (GPCRs). [22] The combination of this peptide-receptor complex can then switch on or off a series of cascading reactions through a multitude of mechanisms. These downstream reactions that are activated may include other G-proteins, tyrosine kinases, and a series of transcription events and thus control all cellular processing and functioning. [22]

In some cases, when a peptide binding to a receptor causing a pathologic "on" state, unregulated transcription and proliferation may ensue, leading to an oncologic state. These cellular processes may be left unchecked and result in tumor growth. The design of synthetic peptides, created to act as endogenous peptides do and bind to a target receptor, can identify the location of tumor growth for identification and even for therapeutic purposes.

Infection

Peptides play a large endogenous role in humans and other species as a first-line barrier to fight infection. One of the components of the body's innate immune system includes the production of antimicrobial peptides (AMPs) in the epithelium. [23] In addition to the epithelium, AMPs are also produced by neutrophils, mast cells, and even adipocytes [24] . These AMPs can be post-translationally modified to fight a wide range of different infections, and with their cationic and interact with the negatively charged bacterial surface. A very important subset of AMPs is called defensins and cathelicidins. [24]

Dermcidin is a known gene that encodes antimicrobial resistance peptides in the sweat glands that can survive at a high salt concentration and over a wide range of pH values. [25] It is known that some bacteria can even produce their AMPs and develop resistance mechanisms to endogenous AMPs so that they can proteolytically cleave the peptides and survive. [24]

Clinical Significance

As previously mentioned, peptides play an essential role in many physiological processes present throughout the body. The clinical significance peptides will be summarized below including some dermatologic disease states as well as therapeutic uses of peptides as imaging probes related to oncology for imaging and tumor targeting. Note that the information provided is concise and is not intended to represent all physiological processes that involve peptides.

Wound repair

Endogenous peptides also play a role in wound healing, and induction of mesenchymal cells to differentiate and promote bacteriolysis within the wound and facilitate healing. Antimicrobial peptides from within wound fluid induced by known as syndecan, a cell surface heparan sulfate proteoglycan. [26] Syndecan functions to activate heparin-binding growth factors and tissue matrix substances to facilitate wound repair in damaged tissues. [26]

Chronic Inflammatory Skin Conditions

As mentioned previously, healthy skin can secrete AMPs to defend against surface attack, especially by gram-positive and gram-negative bacteria, viruses, and fungi. [27] It is shown that peptides play a role have a chronically disrupted epithelial microbiome which predisposes the tissue to pathogenic infection and persistence of inflammatory skin conditions. [28] In the inflamed skin of atopic dermatitis patients compared to the inflamed skin of normal subjects, there is suppression for anti-microbial action due to the decreased expression of normal epidermal AMPs such as LL-37, β-defensin-2, and β-defensin-3. [28]

Where atopic dermatitis patients have suppression of AMPs leading to a further inflammatory, infectious state, patients with rosacea overexpress an anti-microbial peptide known as cathelicidin anti-microbial peptide (CAMP). [27] The metabolites and products of this peptide, CAMP, are what result in the inflammatory state of the epidermis. Knowing the pathway that stimulates CAMP expression in epidermal tissues to elevate anti-microbial peptide generation in an otherwise suppressed state such as atopic dermatitis, may provide a unique and novel therapeutic approach to improve the inflammatory state of patients with this condition. [27]

Molecular Imaging and Tumor Targeting

The mechanism of endogenous peptide and specific receptor binding is principally the design for peptides acting as imaging probes and receptor-binding peptides for overexpressed receptors such as in cancer proliferation. [29] These probes may be strategically designed in vitro to mimic endogenous peptides that ultimately act as biomarkers and allow for the detection of a tumor. The implications of this advanced technology apply to the specific identification of tumor growth and even therapeutic purposes. With advancing sciences and molecular peptide chemistry and a better understanding of more effective targeting, these synthetic peptides have the potential to target multiple disease states with high specificity in imaging modalities such as PET/SPECT, optical imaging, and MRI. [29]

As biochemical sciences and therapeutic design continue to progress, peptide synthesis and design are heavily studied with implications for oncologic therapy as the pharmaceutical industry continues to shift more toward biologicals for new drug candidates. [4]

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Disclosure: Jessica Forbes declares no relevant financial relationships with ineligible companies.

Disclosure: Karthik Krishnamurthy declares no relevant financial relationships with ineligible companies.

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Open access
Published: 18 June 2024

Electrochemical synthesis of peptide aldehydes via C‒N bond cleavage of cyclic amines

Xinyue Fang 1 na1 ,
Yong Zeng 1 na1 ,
Yawen Huang 1 ,
Zile Zhu 2 ,
Shengsheng Lin 1 ,
Wenyan Xu 1 ,
Chengwei Zheng 1 ,
Xinwei Hu 1 ,
Youai Qiu ORCID: orcid.org/0000-0002-5859-6754 2 &
Zhixiong Ruan ORCID: orcid.org/0000-0002-5433-2460 1

Nature Communications volume 15 , Article number: 5181 ( 2024 ) Cite this article

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Electrocatalysis
Synthetic chemistry methodology
Sustainability

Peptide aldehydes are crucial biomolecules essential to various biological systems, driving a continuous demand for efficient synthesis methods. Herein, we develop a metal-free, facile, and biocompatible strategy for direct electrochemical synthesis of unnatural peptide aldehydes. This electro-oxidative approach enabled a step- and atom-economical ring-opening via C‒N bond cleavage, allowing for homoproline-specific peptide diversification and expansion of substrate scope to include amides, esters, and cyclic amines of various sizes. The remarkable efficacy of the electro-synthetic protocol set the stage for the efficient modification and assembly of linear and macrocyclic peptides using a concise synthetic sequence with racemization-free conditions. Moreover, the combination of experiments and density functional theory (DFT) calculations indicates that different N -acyl groups play a decisive role in the reaction activity.

Late-stage peptide C–H alkylation for bioorthogonal C–H activation featuring solid phase peptide synthesis

Complex molecule synthesis by electrocatalytic decarboxylative cross-coupling

Photochemical single-step synthesis of β-amino acid derivatives from alkenes and (hetero)arenes

Introduction.

Peptide aldehydes are a significantly important class of biomolecules, playing a diverse range of essential roles in various living systems 1 . They serve as effective enzyme inhibitors, particularly protease inhibitors, such as Leupeptin 2 , 3 , Elastatinal 4 , and MG101 5 , 6 (Fig. 1A ), thus regulating important biological processes such as digestion, blood clotting, inflammation as well as exhibiting anti-coronavirus activity 7 . Furthermore, aldehydes present in peptides provide a convenient handle for peptide backbone modification or site-specific ligation reactions 8 , 9 , 10 . However, due to the pronounced reactivity of aldehydes, late-stage incorporation in chemical synthesis is required through post-assembly chemical modification, either by employing a specific chemical reaction or by deprotection after peptide elongation to reveal the aldehyde 1 , 11 , 12 , 13 , 14 , 15 , 16 . Therefore, there is an ongoing and strong demand for innovative methods that enable highly efficient synthesis of peptide aldehydes.

A Representative peptide aldehyde biomolecules. B Skeletal diversification of cyclic amines ( C ) Shono-type oxidation. D This work: electro-oxidative ring-opening via C‒N bond cleavage. ABNO = 9-Azabicyclo[3.3.1]nonane N -oxyl.

Organic electrochemistry, utilizing protons and electrons as redox reagents, is emerging as a powerful approach for small-molecule synthesis 17 , 18 , 19 , 20 , 21 , 22 , 23 , 24 , 25 , 26 , 27 , 28 , 29 , 30 . However, it remains underexplored as a tool for achieving mild and chemoselective modification of existing peptides 31 , 32 , 33 . As a mild and customizable reaction platform 34 , electrochemistry offers great promise in overcoming the chemo- and regioselectivity issues encountered in conventional bioconjugation strategies 35 , 36 , 37 , 38 , 39 . Recent contributions in electrochemical modification of peptides were mainly achieved by tyrosine- 40 , 41 , 42 , 43 , 44 , 45 and tryptophan- 46 , 47 specific functionalization as well as side chain diversification via direct or indirect electrochemical methods. Recognizing the necessity for additional research methodologies to facilitate the coupling of specific functional groups, various structural functionalization reactions of cyclic amines 48 were initiated, in order to access distinctive chemical and functional spaces. These reactions encompassed ring-opening reactions 49 , 50 , 51 , ring-expansion reactions 52 , ring contraction 53 , heterocycle replacement 54 , and C2-direct functionalization reactions 55 , 56 . These innovative approaches involved the breaking of conventional saturated C‒C or C‒N bonds for chemical bond reorganization (Fig. 1B ). In addition, with regard to piperidine derivatives, prevalent in pesticides and peptide-based pharmaceuticals 57 , 58 , there is a wealth of examples employing the Shono-type oxidation strategy to achieve C2 functionalization of piperidines (Fig. 1C ) 59 , 60 , 61 . However, there are few reports on electrochemical C‒N bond ring-opening reactions. Within our program on sustainable electrochemical modification of peptides 62 , 63 and cyclic amines 64 , 65 , herein we present an electro-oxidative ring-opening approach to achieve unnatural peptide aldehydes (Fig. 1D ). Notable features of our general strategy include (i) (homo)proline-specific diversification of peptide backbones using exceedingly mild and biocompatible conditions in a metal-, chemical oxidant-, and racemization-free fashion, (ii) a broad substrate scope with various peptides and complex bioactive molecules, (iii) step- and atom- economical ring-opening procedure for smoothly yielding peptide aldehydes and remote amino aldehydes via selective C‒N bond cleavage, and (iv) setting the stage for versatile syntheses of macrocyclic peptides. Detailed mechanistic insights of this process provided strong support for the presence and pivotal role of an N -Piv protecting group.

Optimization of reaction conditions

We commenced our investigations by probing a variety of different electrolysis conditions, employing an undivided cell equipped with a graphite felt anode and a platinum cathode, for the envisioned electro-oxidative ring-opening of D -homoproline-containing dipeptide (Piv-Homopro-Leu-OMe) 1a . The optimal results were obtained when dipeptide 1a was directly electrolyzed at a constant current of 8.0 mA in a mixed electrolyte solution of n Bu 4 NHSO 4 in MeCN/H 2 O (9:1) at room temperature under atmospheric conditions without additional oxidants and transition metal catalysts. Under these conditions, the desired dipeptide aldehyde 1b was isolated in 73% yield without racemization (Table 1 , entry 1) (For details, please see the Supplementary Table 1 ). The electrolyte and solvent were both found to be critical factors for the reaction to achieve the optimal yield (entries 2‒6). Among a variety of electrolytes, n Bu 4 NPF 6 and n Bu 4 NOAc and n Bu 4 NOH showed poor efficacy (entries 2‒4), while LiClO 4 , n Bu 4 NI, or no electrolyte displayed inactive performance, highlighting the critical importance of HSO 4 ¯ (entries 5‒6), which also confirmed by the further DFT calculations. In addition, solvent screening experiments verified the essential role of H 2 O as the oxygen source, since performing the electrolysis in the solvent with traceless H 2 O produced the remote amino aldehyde 1b in extremely low yields (entries 7‒9). The choice of electrode material proved critical because a dramatically reduced product yield was obtained when using other types of carbon electrodes (entries 10‒12). A further control experiment indicated that electricity was indispensable to promote the desired reaction (entry 13).

Substrate scope

With the optimal conditions in hand, we explored the scope of the electro-oxidative C‒N bond cleavage protocol (Fig. 2 ). The electrochemical ring-opening reaction exhibited excellent compatibility with a variety of natural aliphatic amino acids such as leucine (Leu), alanine (Ala), valine (Val), threonine (Thr), glycine (Gly), and glutamic acid (Glu) as well as unnatural medicinally-useful bulky amino acids, e.g., L- tert -leucine, L-allylglycine, L-cyclohexylglycine, and cycloleucine, affording the dipeptide aldehydes in moderate to good yields ( 1b – 14b ). The structure of 11b was unambiguously confirmed by single-crystal X-ray diffraction studies. Thereafter, we investigated the scope of tripeptides conjugated with a diversity of amino acid residues ( 15b – 22b ). The robust nature of the electro-oxidative ring-opening transformation was reflected by fully tolerating a wealth of valuable transformation groups, including alkenes and alkynes, which could serve as a handle for future late-stage bioconjugation. More importantly, the peptides containing electron-rich phenylalanine residues were compatible under slightly modified electrolytic conditions, which normally readily decomposed under direct electrochemical conditions. To our delight, the strategy for the direct electrochemical synthesis of peptide aldehyde smoothly provided L-allylglycine-containing tetrapeptides and bulky alkyl group-containing pentapeptides, further emphasizing the strong biocompatible conditions ( 23b – 25b ). Furthermore, a wide range of common amide and ester substrates with bulky ( iso -propyl, tert -butyl, cyclohexyl, adamantyl) or linear alkyl groups ( n -butyl), including electron-donating (-OMe) and electron-withdrawing substituents (-CO 2 Me, -CF 3 , -CN), as well as synthetic useful handles (alkenyl, alkynyl, linker), were found to be fully tolerated by the optimized electrooxidation ( 26b – 44b ). Gratifyingly, the practical utility of our approach was further illustrated by successfully performing the desired ring-opening with substrates derived from marketed drug pregabalin, which effectively delivered ε -amino aldehyde derivative 35b in moderate yield. It is noteworthy that the antioxidant activity of aldehyde products was evaluated by DPPH (2,2-Diphenyl-1-picrylhydrazyl) radical-scavenging activity assays, some peptides, such as 21b, 22b, and 24b , exhibited mild antioxidant activity at a concentration-dependent manner (For details, please see the Supplementary Figs. 14 and 15 ).

Reaction conditions: Undivided cell, graphite felt (GF) anode, platinum plate (Pt) cathode, 1a-44a (0.3 mmol), n Bu 4 NHSO 4 (0.3 mmol), MeCN/H 2 O (9:1, 10 mL), constant current = 8.0 mA, 6 h (6.0 F ), RT, under air. Isolated yields are reported. [a] 9.0 h. [b] Yields based on recovered starting materials. [c] ketoABNO (50 mol %).

Furthermore, encouraged by these exciting results, we sought to investigate the synthetic applications of the peptide aldehyde product by functional group transformation (Fig. 3 ). Thus, the outstanding potential of our ring-opening approach was demonstrated by its easy scalability and versatile transformations of the generated peptide aldehyde. For instance, we electrolyzed 5.0 mmol of dipeptide 1a to deliver the corresponding peptide aldehyde 1b in 72% yield, without appreciable loss in efficacy. In addition, five novel peptide sequences ( 45 – 49 ) were each rapidly constructed in simple steps from 1b , highlighting the potential for such reactions to enable efficient diversification of native residues in preassembled peptide building blocks, including the synthesis of unnatural amino acids. Strikingly, a series of peptide-conjugated drugs ( 50 – 56 ) were further assembled through ligation with marketed drugs via amination or esterification.

Isolated yields are reported. [a] NaBH 4 , MeOH, 0 °C, 1 h. [b] (i) NaBH 4 , MeOH, 0 °C, 1 h; (ii) DAST, DCM, −78 °C to RT, 14 h. [c] (i) NaBH 4 , MeOH, 0 °C, 1 h; (ii) MsCl, Et 3 N, DCM, 0 °C to RT, 4 h; (iii) NaN 3 , DMF, RT, 12 h. [d] PPh 3 MeBr, t -BuONa, THF, 0 °C to RT, 12 h. [e] O 2 , NHPI, MeCN, RT, 12 h. [f] Transformations of hydroxyl peptides. [g] Applications of acid products. For details, please see the Supplementary Information on pages 57−69.

In the realm of non-ribosomal peptide synthetase natural products, macrocyclic peptides stand out as highly prized therapeutic contenders compared to their linear counterparts. This preference is rooted in their heightened resistance to chemical and enzymatic degradation, improved receptor specificity, and advantageous pharmacokinetic profiles 49 , 66 , 67 , 68 , 69 . We sought to investigate the feasibility of efficiently constructing and expanding macrocycles by rapidly introducing new functional groups into peptides starting from a simple homoproline residue. Gratifyingly, olefin- and carboxylic acid-derived dipeptides ( 48 and 49 ) as well as tripeptide aldehyde ( 22b ) could be rapidly transformed into four macrocycles containing ( E )-alkene ( 58 ), amide ( 59 ), ester and alkyne ( 60 ) 70 , as well as triazole ( 61 ) linkers, respectively, through concise synthetic sequences by key olefin metathesis, manganese-catalyzed C–H alkynylation, and click reaction steps (Fig. 4 ) (For details, please see the Supplementary Information on pages 69−80). The incorporation of functional groups into the linker region of stapled peptide-like structures has been demonstrated to influence the biological properties of the resulting products. Our post-assembly electro-oxidative strategy enables the rapid synthesis of a diverse array of functionally and structurally enriched molecules, as exemplified by the small library of compounds synthesized from dipeptide aldehyde 1b .

The synthesis of macrocyclic peptides 58–61.

Remote amino aldehydes, especially unnatural derivatives, serving as direct precursors of variant amino acids, are indispensable building blocks in the synthesis of peptide aldehydes 1 , 71 , 72 . Thus, the scope of cyclic amines 73 , 74 , 75 was also investigated using a slightly modified set of conditions (Fig. 5 and Supplementary Table 2 in Supplementary Information). Diverse substitution patterns on the piperidine ring exhibited excellent tolerance, enabling the synthesis of the corresponding acyclic amino aldehydes with moderate to good yields (up to 88%). Notably, benzoyl-protected piperidine afforded the corresponding aldehyde in moderate yield ( 62b ), while that with other protected groups Boc and Cbz resulted in ortho -hydroxylation products instead of the desired aldehydes (For details, please see the Supplementary Table 3 ). Piperidines with ester functional groups on the saturated azacyclic backbone ( 66a and 68a ) exhibited effective performance. However, cyclic amines with free carboxylic acid groups at the α-positions ( 69a and 70a ) produced the respective decarboxylated amino aldehydes. Piperidines containing benzylic sites prone to oxidation yielded the corresponding aldehydes with a 57% yield ( 65b ). Remarkably, complete positional selectivity was observed with 2-substituted azacycles ( 64a, 66a , and 68a ) in the oxidative ring-opening protocol. The scalability of our ring-opening approach was demonstrated by a successful gram-scale reaction ( 63b ). The method also tolerated saturated azacycles of various ring sizes (four to eight-membered rings), resulting in β -, γ -, or remote amino aldehydes ( 67b, 72b , and 73b ), although the isolated yield for β -amino aldehyde ( 71b ) was low. Moreover, the α,β -unsaturated aldehyde 74b was obtained when the piperidine’s para-position was substituted with a methoxy group. Finally, both substituted cyclic amines 75a and 76a could be converted into corresponding ring-opening products, although the chemo-selectivity was unromantic.

Reaction conditions: Undivided cell, graphite felt (GF) anode, platinum plate (Pt) cathode, 62a-76a (0.3 mmol), n Bu 4 NPF 6 (0.3 mmol), MeCN/H 2 O (9:1, 10 mL), constant current = 8.0 mA, 3 h (3.0 F ), RT, under air. Isolated yields are reported. [a] 6.0 h. [b] Yield based on recovered starting materials.

Mechanistic studies

In light of the outstanding versatility of the electrochemical ring-opening methodology, we were intrigued to delineate its mode of action. To this end, an isotopic labeling experiment was conducted in the presence of H 2 18 O under the standard conditions. The results indicated that the oxygen atom in the newly formed aldehyde derived from the isotopically labeled water (Fig. 6A ). Experiments with isotopically labeled co-solvents unraveled a facile C–H activation step, as evidenced by H/D scrambling at the α -position of the aldehyde (Fig. 6B ). Furthermore, analysis of the voltammogram results disclosed that the dipeptide 1a underwent anodic oxidation at ~1.928 V (vs Ag/AgCl), while no distinct oxidation peak was observed for 1b , indicating the difficulty of further oxidizing aldehyde 1b to acid 49 (Fig. 6C ). This observation provided a plausible explanation for the absence of carboxylic acid products in the electrolytic reaction, even when using higher currents. Additionally, an electricity on/off study revealed that any chain processes were transient and short-lived (Fig. 6D ). This finding suggested that electrolysis is required for sustained product formation, indicating that the reaction was less likely to proceed through a radical chain mechanism.

A Isotopic labeling experiment. B H/D exchange experiment. C Cyclic voltammograms. D Electricity on/off experiment.

Furthermore, DFT calculations were performed to further elucidate the reaction mechanism using 63a as a model substrate (see Fig. 7 and Supplementary Table 7 in Supplementary Information). As is shown in Fig. 7 , 63a may first undergo two single electron anodic oxidation and the iminium intermediate Int3 emerges. According to the potential energy surface, across TS2 , water capture of Int3 is endothermic (Δ G = 8.98, Δ G * = 14.70). In the yielding Int5 , an O \(-\) H···O intramolecular hydrogen bond is found to stabilize the intermediate, while the analogous intermediate from water capture of Int3′ cannot be located. Taking HSO 4 – in n Bu 4 NHSO 4 as a Bronsted base, the resulting Int5 may deprotonate barrierlessly into Int6 , and reprotonate into Int7 via TS3 , dissipating 3.32 kcal/mol Gibbs free energy. The proton transferring is a fast process due to low energy barriers. A C \(-\) N bond cleavage will take place in Int7 , mounting an energy barrier of 12.12 kcal/mol via TS4 , yielding protonated product Int8 , which may associate with HSO 4 – into Int9 . Moreover, another reaction pathway was also considered, in which the C \(-\) N bond cleavage takes place in Int6 via TS5 yielding an enol-like product. However, the energy barrier (26.72 kcal/mol) is too high to cross.

All energy units are kcal/mol.

We then altered the N -acyl group into Boc, Bz, and Cbz, and calculated the relative Gibbs free energy of some corresponding intermediates and transition states. Results are listed in Table 2 . In each variation, Int3 is selected as the energy zero-point.

With DFT results in hands, the insight into reactivity of substrates can be gained. According to the proposed mechanism, applying the steady-state approximation and Eyring equation, a rate equation in proportion to the concentration of iminium ( Int3 ) could be derived (To check the process of deduction, please see the Supplementary Information on pages 91 − 93):

Where r is the reaction rate of Int3 ’s conversion to the product form, k obs denotes the observed rate constant, k B is the Boltzmann constant, T = 298.15 K, h is the Planck constant, R is the gas constant, [H 2 O] is the concentration of water, and c (Int3) is the analytic concentration of Int3 , the iminium intermediate. According to the formula (Eq. 2 ), for each variation of the N -acyl group, the free energy difference between TS4 and Int3 plays a pivotal role in deciding the reactivity. Using the DFT-calculated data from Table 2 , different N -acyl groups bear different k obs calculated in Table 3 . Among Boc, Bz, Cbz, and Piv-substituted cyclic amines, the Piv-substituted reacts the fastest (8.9 × 10 –2 L mol –1 s –1 ), but with an oxygen atom inserted (the Boc-substituted), k obs drops prominently to 7.2 × 10 –7 L mol –1 s –1 , which implies the scarce reactivity.

In summary, we have reported a pioneering electrochemical ring-opening protocol that enables direct synthesis of unnatural peptide aldehydes and remote amino aldehydes through mild and biocompatible reaction conditions involving C‒N bond cleavage. The strategy offers expedient access to a wide variety of peptide aldehydes with potential antioxidant activities, as well as expanded substrate scope including amides, esters, and cyclic amines with diverse synthetic functional groups. The unique power of the electro-oxidative ring-opening approach set the stage for efficient modification and assembly of acyclic and macrocyclic peptides by short synthetic sequence under racemization-free conditions. Notably, the experiments and DFT calculations demonstrate that different N -acyl groups play a decisive role for the reaction.

General procedure for electrochemical reactions

In an undivided cell (30 mL) equipped with a stirring bar, a mixture of substrates (0.3 mmol), n Bu 4 NHSO 4 (0.3 mmol, 0.03 M), and MeCN/H 2 O (9:1, 10 mL) were added. The cell was equipped with graphite felt plate (GF, 1.5 cm × 1.0 cm × 0.2 cm) as the anode and platinum plate (Pt, 1.5 cm × 1.0 cm × 0.01 cm) as the cathode connected to an AXIOMET AX-3003P DC regulated power supply. The reaction mixture was stirred and electrolyzed at a constant current of 8 mA at room temperature for 3~6 h. Upon completion, the solvent was removed directly under reduced pressure to afford the crude product, which was further purified by flash column chromatography to afford the desired products.

Data availability

The authors declare that the data supporting the findings of this study are available within the paper and its Supplementary Information. Extra data are available from the corresponding author upon request. Source data are provided with this paper. Crystallographic data for the structures reported in this Article have been deposited at the Cambridge Crystallographic Data Centre, under deposition numbers CCDC 2216047 (11b). These data can be obtained free of charge from The Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/data_request/cif . Source data are provided with this paper.

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Acknowledgements

Support by the National Natural Science Foundation of China (22271067 to Z.R., 22201052 to X.H.), Key-Area Research Project of Guangdong Provincial Department of Education (2022ZDZX2051 to Z.R.), Guangzhou Science and Technology Project (2023A04J0696 to X.H.), and the Plan on Enhancing Scientific Research in Guangzhou Medical University (GMU) (Z.R.) is most gratefully acknowledged.

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These authors contributed equally: Xinyue Fang, Yong Zeng.

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Guangzhou Municipal and Guangdong Provincial Key Laboratory of Molecular Target & Clinical Pharmacology and the State Key Laboratory of Respiratory Disease, School of Pharmaceutical Sciences & the Fifth Affiliated Hospital, Guangzhou Medical University, Guangzhou, 511436, PR China

Xinyue Fang, Yong Zeng, Yawen Huang, Shengsheng Lin, Wenyan Xu, Chengwei Zheng, Xinwei Hu & Zhixiong Ruan

State Key Laboratory and Institute of Elemento-Organic Chemistry, Frontiers Science Center for New Organic Matter, College of Chemistry, Nankai University, 94 Weijin Road, Tianjin, 300071, PR China

Zile Zhu & Youai Qiu

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Z.R. directed the project and wrote the manuscript. Z.R., Y.Q., X.H., and X.F. conceived and designed the study and wrote the draft manuscript. X.F., Y.Z., Y.H., S.L., and C.Z. performed the experiments, mechanistic studies and analyzed the data. Y.Q. and Z.Z. performed the DFT calculations and analyzed the data. W.X. performed the bioactive assay. All authors contributed to scientific discussion.

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Correspondence to Xinwei Hu , Youai Qiu or Zhixiong Ruan .

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Fang, X., Zeng, Y., Huang, Y. et al. Electrochemical synthesis of peptide aldehydes via C‒N bond cleavage of cyclic amines. Nat Commun 15 , 5181 (2024). https://doi.org/10.1038/s41467-024-49223-y

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Fundamentals of Modern Peptide Synthesis

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Muriel Amblard 2 ,
Jean-Alain Fehrentz 2 ,
Jean Martinez 2 &
Gilles Subra 2

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 298))

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The purpose of this chapter is to delineate strategic considerations and provide practical procedures to enable non-experts to synthesize peptides with a reasonable chance of success. This chapter focuses on Fmoc chemistry, which is now the most commonly employed strategy for solid phase peptide synthesis (SPPS). Protocols for the synthesis of fully deprotected peptides are presented, together with a review of linkers and supports currently employed for SPPS. The principles and the different steps of SPPS (anchoring, deprotection, coupling reaction, and cleavage) are all discussed, along with their possible side reactions.

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Peptide Synthesis: Methods and Protocols

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Amblard, M., Fehrentz, JA., Martinez, J., Subra, G. (2005). Fundamentals of Modern Peptide Synthesis. In: Howl, J. (eds) Peptide Synthesis and Applications. Methods in Molecular Biology™, vol 298. Humana Press. https://doi.org/10.1385/1-59259-877-3:003

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Benzeneseleninic acid used as an oxidizing and deprotecting reagent for the synthesis of multi-cyclic peptides constrained by multiple disulfide bonds and thioether bridges †.

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a College of Chemistry and Materials Science, Key Laboratory of Analytical Science and Technology of Hebei Province, and MOE Key Laboratory of Medicinal Chemistry and Molecular Diagnostics, Hebei University, Baoding 071002, Hebei Province, China E-mail: [email protected] , [email protected]

b College of Chemistry, Chemical Engineering and Materials Science, Zaozhuang University, Zaozhuang 277160, Shandong Province, China E-mail: [email protected]

Peptides constrained by multiple disulfide bonds (MDBs) or by thioether bridges play a critical role in improving the biological and pharmaceutical activities of peptide drugs. The synthesis of correct MDBs and thioether-bridged bicyclic peptides still remains a significant challenge. In this work, benzeneseleninic acid (BSA) acting as both an oxidant and a deprotecting reagent for the synthesis of MDB- and thioether-bridged bicyclic peptides has been investigated. Disulfide bonds in peptides can be formed by direct oxidation of two sulfhydryl groups by BSA in neutral media or via two concerted steps, namely deprotecting two acetamidomethyl (Acm) groups and oxidation by BSA in acidic media. As such, two disulfide bonds in α-conotoxin SI, apamin, α-conotoxin IMI and a peptide containing a methionine residue were synthesized regioselectively by the use of the BSA oxidation and deprotection reaction (BSA-ODr). By utilization of the BSA deprotection property, bicyclic peptides were synthesized based on the crosslinking of xylylene dibromide with sulfhydryl groups. Furthermore, two disulfide bonds in α-conotoxin SI and three disulfide bonds in conotoxin mr3e, enterotoxin STp, μ-conotoxin KIIIA, linaclotide and ziconotide were also synthesized regioselectively through oxidation of fully reduced peptides by BSA. All the reactions were carried out under mild conditions in a one-pot manner and peptides with satisfactory yields were achieved. In addition, the BSA-ODr is compatible with methionine residues in the peptides. Moreover, the relative positions of two Acm-protected cysteines and two free cysteines have no impact on the BSA-ODr approach for the construction of two disulfide bonds in peptides. The oxidative folding strategies based on BSA can be executed in peptide manufacture. BSA is readily accessible conferring efficient BSA-ODr and oxidative folding methodologies for the synthesis of MBDs and thioether bridges in peptides.

Graphical abstract: Benzeneseleninic acid used as an oxidizing and deprotecting reagent for the synthesis of multi-cyclic peptides constrained by multiple disulfide bonds and thioether bridges

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Benzeneseleninic acid used as an oxidizing and deprotecting reagent for the synthesis of multi-cyclic peptides constrained by multiple disulfide bonds and thioether bridges

Y. Xing, T. Bo, N. Zhang, M. Wu, J. Wang, S. Shen, Y. Wang, C. Song, T. Shi and S. Huo, Org. Chem. Front. , 2024, Advance Article , DOI: 10.1039/D4QO00589A

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Synthesis and evaluation of the first 68 ga-labeled c -terminal hydroxamate-derived gastrin-releasing peptide receptor-targeted tracers for cancer imaging with positron emission tomography.

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Wang, L.; Kuo, H.-T.; Chen, C.-C.; Chapple, D.; Colpo, N.; Ng, P.; Lau, W.S.; Jozi, S.; Bénard, F.; Lin, K.-S. Synthesis and Evaluation of the First 68 Ga-Labeled C -Terminal Hydroxamate-Derived Gastrin-Releasing Peptide Receptor-Targeted Tracers for Cancer Imaging with Positron Emission Tomography. Molecules 2024 , 29 , 3102. https://doi.org/10.3390/molecules29133102

Wang L, Kuo H-T, Chen C-C, Chapple D, Colpo N, Ng P, Lau WS, Jozi S, Bénard F, Lin K-S. Synthesis and Evaluation of the First 68 Ga-Labeled C -Terminal Hydroxamate-Derived Gastrin-Releasing Peptide Receptor-Targeted Tracers for Cancer Imaging with Positron Emission Tomography. Molecules . 2024; 29(13):3102. https://doi.org/10.3390/molecules29133102

Wang, Lei, Hsiou-Ting Kuo, Chao-Cheng Chen, Devon Chapple, Nadine Colpo, Pauline Ng, Wing Sum Lau, Shireen Jozi, François Bénard, and Kuo-Shyan Lin. 2024. "Synthesis and Evaluation of the First 68 Ga-Labeled C -Terminal Hydroxamate-Derived Gastrin-Releasing Peptide Receptor-Targeted Tracers for Cancer Imaging with Positron Emission Tomography" Molecules 29, no. 13: 3102. https://doi.org/10.3390/molecules29133102

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Peptide Synthesis
Peptide Synthesis
Peptide Synthesis
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Peptide Synthesis and Characterization Stages

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PEPTIDE SYNTHESIS || CLASSICAL PEPTIDE SYNTHESIS AND ITS DRAWBACKS
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SOLID AND SOLUTION PHASE PEPTIDE SYNTHESIS BSC FINAL YEAR

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Peptides Guide
Learn about peptides, their structure, functions and importance in biology and medicine. Find out how peptides are synthesized using solution-phase and solid-phase methods, and explore examples of peptide drugs and research tools.
Peptide Synthesis
In solid-phase peptide synthesis (SPPS) one peptide end is attached to water-insoluble polymer and remains protected throughout the entire peptide formation, meaning both fewer steps and simplified purification, the reagents can be rinsed away without losing the peptide. Solid-phase peptide synthesis involves the following steps: attaching ...
Peptide synthesis
In organic chemistry, peptide synthesis is the production of peptides, compounds where multiple amino acids are linked via amide bonds, also known as peptide bonds. Peptides are chemically synthesized by the condensation reaction of the carboxyl group of one amino acid to the amino group of another. Protecting group strategies are usually ...
Introduction to Peptide Synthesis
The concept of solid-phase peptide synthesis (SPPS) is to retain chemistry that has been proven in solution but to add a covalent attachment step that links the nascent peptide chain to an insoluble polymeric support (resin). Subsequently, the anchored peptide is extended by a series of addition cycles ( Fig. 18.1.1 ).
26.8: Peptide Synthesis
Once the structure of a peptide is known, its synthesis can be undertaken—perhaps to obtain a larger amount for biological evaluation. A simple amide might be formed by treating an amine and a carboxylic acid with a carbodiimide (either DCC or EDC; Section 21.3), but peptide synthesis is a more difficult problem because many different amide bonds must be formed in a specific order, rather ...
Therapeutic peptides: current applications and future directions
The chemical synthesis of peptides is well-developed, particularly solid-phase peptide synthesis (SPPS) technology developed by Merrifield in 1963 114. SPPS technology has since been remarkably ...
Bioactive Peptides: Synthesis, Sources, Applications, and Proposed
Chemical synthesis of peptides by soluble-phase methods was first used in 1953 to produce pharmaceutical insulin peptides . The basis of this method is the reaction of amino acids in a soluble medium. Perhaps the most important advantage of soluble-phase methods is the economic justification and purification at each stage of the synthesis since ...
Peptide Synthesis: Methods and Protocols
A chapter from a book series on peptide therapeutics that covers the history, fundamentals, and methods of peptide synthesis. Learn about the protecting groups, resins, linkers, and coupling reactions used in solution-phase and solid-phase peptide synthesis.
Peptides: Molecular Basis, Secondary Structures, and Synthesis Methods
Peptides are omnipresent in nature and indispensable for life activities. Inspired by nature, human body regulates peptide molecular structure and uses peptide self-assembly to achieve various applications, which rely on the synthesis of peptide molecular compounds.
Peptide synthesis: Methods, trends, and challenges
Peptide scientists still have a number of synthetic challenges to address including the synthesis of long peptides, bicyclic peptides, pegylated peptides, lipopeptides, glycosylated peptides, cell-penetrating peptides, and peptide-peptoid hybrids. The "Peptide Era" just started. Recommended articles.
Peptide Synthesis: Methods and Protocols
A book series that provides step-by-step protocols for synthesizing peptides and their derivatives for various applications. Learn about cutting-edge techniques, key implementation advice, and troubleshooting tips from the experts.
Peptide Synthesis
Peptide synthesis is characterized as the formation of a peptide bond between two amino acids. While there is no definitive definition of a peptide, it usually refers to flexible (little secondary structure) chains of up to 30-50 amino acids. The ability to form peptide bonds to link amino acids together is over 100 years old, although the ...
26.7 Peptide Synthesis
26.7 • Peptide Synthesis Once the structure of a peptide is known, its synthesis can be undertaken—perhaps to obtain a larger amount for biological evaluation. A simple amide might be formed by treating an amine and a carboxylic acid with a carbodiimide (either DCC or EDC; ...
Synthesis of Peptides
What Are Peptide Bonds? A "peptide bond" is an amide linkage (see Amides: Properties. Synthesis, and Nomenclature) that connects two amino acids, as in the "dipeptides" L-phenylalanyl-L-valine (below left) and L-leucyl-L-alanine (below right): 2. The "Proteinogenic" Amino Acids. Proteinogenic amino acids are the building blocks of ...
Biochemistry, Peptide
Peptide synthesis depends on three main reactions: 1. an amino acid goes through a deprotection step, a preparatory reaction that adds the next amino acid to the chain, and lastly, a coupling reaction that forms the final peptide with functionality. In the second step, the amino acid becomes activated with several reagents.
Solid-phase peptide synthesis: from standard procedures to the ...
Peptides play a pivotal role in biological, medical and pharmaceutical research. Therefore, the synthesis of such polyamide structures has been a major focus of organic chemistry for over a century.
Solid-Phase Peptide Synthesis: An Introduction
Learn the basics of peptide chemistry and solid-phase peptide synthesis (SPPS) from this chapter in a book series on methods in molecular biology. It covers the history, reagents, mechanisms, and applications of SPPS, as well as solution synthesis and chemical ligation.
26.8: Peptide Synthesis
The strategy for peptide synthesis, as outlined here, should now be apparent. The following example shows a selective synthesis of the dipeptide Ala-Gly. An important issue remains to be addressed. Since the N-protective group is an amide, removal of this function might require conditions that would also cleave the just formed peptide bond.
24.8: Peptide Synthesis
The strategy for peptide synthesis, as outlined here, should now be apparent. The following example shows a selective synthesis of the dipeptide Ala-Gly. An important issue remains to be addressed. Since the N-protective group is an amide, removal of this function might require conditions that would also cleave the just formed peptide bond.
Peptide Synthesis
What is Peptide Synthesis? Characterized by the formation of a peptide bond between two amino acids, peptide synthesis is, essentially, the production of peptides. Though peptide synthesis was somewhat hampered by relatively inefficient production practices at its inception, advancements in chemistry and technology have led to vastly improved synthesis methods. With the strong growth of …
Electrochemical synthesis of peptide aldehydes via C‒N bond ...
Peptide aldehydes are an important class of biomolecules, playing essential roles in various living systems, and there is an ongoing demand for the efficient synthesis of peptide aldehydes. Herein ...
Ru(II)-Catalyzed C(sp2)-H Activation Annulation: Synthesis of
Benzoisoquinolones are aryl ring extended isoquinolinone derivatives, which are constituents of alkaloid natural products. This report describes the synthesis of novel benzoisoquinolone amino acid/peptide derivatives from the respective N-aryl amino esters/peptides through Ru-catalyzed C(sp 2)-H annulation at room temperature.The N-terminal amide acts as an intrinsic directing group and ...
Fundamentals of Modern Peptide Synthesis
Learn how to synthesize peptides using solid phase peptide synthesis (SPPS) with Fmoc/tBu strategy. This chapter covers the principles, materials, methods, and side reactions of SPPS, with protocols and examples.
Benzeneseleninic acid used as an oxidizing and deprotecting reagent for
Peptides constrained by multiple disulfide bonds (MDBs) or by thioether bridges play a critical role in improving the biological and pharmaceutical activities of peptide drugs. The synthesis of correct MDBs and thioether-bridged bicyclic peptides still remains a significant challenge. In this work, benzeneseleninic
Molecules
Gastrin-releasing peptide receptor (GRPR), overexpressed in many solid tumors, is a promising imaging marker and therapeutic target. ... "Synthesis and Evaluation of the First 68 Ga-Labeled C-Terminal Hydroxamate-Derived Gastrin-Releasing Peptide Receptor-Targeted Tracers for Cancer Imaging with Positron Emission Tomography" Molecules 29, no ...
25.8: Peptide Synthesis
The strategy for peptide synthesis, as outlined here, should now be apparent. The following example shows a selective synthesis of the dipeptide Ala-Gly. An important issue remains to be addressed. Since the N-protective group is an amide, removal of this function might require conditions that would also cleave the just formed peptide bond.

Peptide Synthesis

Page contents

Process of synthesizing peptides

View and select products

Introduction to peptide synthesis

Select products

Peptide deprotection

Amino acid coupling

Peptide cleavage

26.7 Peptide Synthesis

(Advanced) References and Further Reading

2. The “Proteinogenic” Amino Acids

3. Synthesis of A Simple Dipeptide Without Protecting Groups (is not advisable!)

4. Synthesis Of A Dipeptide Using A Protecting Group Strategy

5. Synthesis of Longer Peptides – Tripeptides and Tetrapeptides

6. Bonus Topic: Solid-Phase Peptide Synthesis

00 General Chemistry Review

01 Bonding, Structure, and Resonance

02 Acid Base Reactions

03 Alkanes and Nomenclature

04 Conformations and Cycloalkanes

05 A Primer On Organic Reactions

06 Free Radical Reactions

07 Stereochemistry and Chirality

08 Substitution Reactions

09 Elimination Reactions

10 Rearrangements

11 SN1/SN2/E1/E2 Decision

12 Alkene Reactions

13 Alkyne Reactions

14 Alcohols, Epoxides and Ethers

15 Organometallics

16 Spectroscopy

17 Dienes and MO Theory

18 Aromaticity

19 Reactions of Aromatic Molecules

20 Aldehydes and Ketones

21 Carboxylic Acid Derivatives

22 Enols and Enolates

24 Carbohydrates

25 Fun and Miscellaneous

26 Organic Chemistry Tips and Tricks

27 Case Studies of Successful O-Chem Students

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