In 1962, Merrifield established the solid-phase peptide synthesis method, and since then, solid-phase synthesis using protected amino acids as raw materials has attracted people's attention due to its unique advantages. 

In the following 10 years, the synthesis of peptides in the same phase continued to improve, developing from small-scale, short peptide chain synthesis to large-scale, long peptide chain synthesis. The anti AIDS drug T-20, which was launched in 2003, is a fully synthesized small peptide drug composed of 36 amino acid residues.

In the 1990s, after the combination of combinatorial chemistry and peptide synthesis technology, the synthesis of combinatorial peptide libraries appeared, which played a huge role in promoting the fields of chemistry, biochemistry, medicine, molecular biology, etc. In the process of development, the requirements for protecting amino acids have also been constantly increasing, leading to continuous improvement.

Solid-phase peptide synthesis Method

In solid-phase peptide synthesis, it is required that the active functional groups of the amino acids involved in the reaction that do not participate in the formation of peptide bonds are protected. At the same time, after the reaction is completed, the protective groups of these active groups can be removed.

When selecting amino acid protection groups, it is necessary to fully consider the issue of reaction selectivity, namely α -  Amino protective groups and α -  The carboxyl protective group is a temporary protective group, and once an amino acid binds to the peptide chain, it must be able to be immediately removed; The side chain protection group is a "permanent" protection group that is not affected by the reactants during the peptide synthesis process and does not react. It is finally removed after the peptide synthesis is completed. So the stability of the side chain protection group is greater than α -  Amino protective groups and α -  Carboxyl protective groups.

The selection of protective amino acids is directly related to the yield and purity of synthesized products. The by-products and racemization formed during the reaction pose great difficulties for purification. Therefore, developing new types of protective amino acids is an important component of solid-phase peptide synthesis.

At present, amino acid protection is mainly divided into three categories: α -  Amino protection α-  Carboxyl protection and side chain active group protection. Based on these three classifications, this article focuses on introducing some new varieties of protective amino acids in recent years, their structural characteristics, and application conditions.

α -  Amino protection
In principle, amino groups can undergo reversible shielding through reactions such as acylation, alkylation, and alkylation. Protective groups based on sulfur and phosphorus derivatives have also been reported.
Over the past decade, hundreds of different amino protection groups have been developed, mainly including alkoxycarbonyl protection groups, acyl protection groups, and alkyl protection groups. The reported research results indicate that there is no universal ideal amino protection group for solid-phase synthesis of peptides.

α -  Carboxyl protection
Solid phase synthesis of peptides - generally not required α -  The protection of carboxyl groups is only necessary under very few conditions α -  Carboxyl group. If synthesizing glutathione, it requires glutamic acid γ -  Carboxyl groups react, while α-  Carboxyl groups require protection. At present, hydroxyl protection groups are roughly divided into two categories: ester groups and hydrazides.

Protection of side chain active groups
ω -  The carboxyl protecting groups Asp and G1u have two carboxyl groups, which, under the action of a base or strong acid, are on the side chain ω -  Carboxyl groups are prone to side reactions. For example, the side chain ester formed during the activation of aspartic acid will generate iso asparagine peptide through the corresponding intermediate state of succinimide.

Commonly used protective groups include tert butyl ester group (OB) used in the F strategy and tert butyl ester group used in the Boc strategy β -  Cyclohexyl ester group (OcHex). These two protective groups can usually inhibit the occurrence of the aforementioned side reactions.

However, when Asp is connected to Gly, Thr (tBu), Cys (Acm), the side reactions are very severe, and N-2-hydroxy-4-methoxyphenyl (Hmb) protection must be used. When removing the protective group, TFA treatment can be used to add mercaptan to capture carbon cations, reducing the occurrence of side reactions.

Protecting amino acids is a key raw material for peptide synthesis, and the strategies for protection vary depending on the method of peptide synthesis. At present, several companies have produced commercialized protective amino acids and peptides, such as Nova, Aldrich Sigma, PE, Omizzur ltd, etc.

The development focus of protecting amino acids is mainly focused on: ① the development of new types of amino acids α-  The development of amino protection groups, Nsc, Dde, Nde and other groups have the characteristics of reducing racemization and improving deprotection efficiency; ② For the development of side chain active group protection, various protective groups with excellent stability, high selectivity, and reduced side reactions have been developed for different active groups.

The protective amino acids prepared in practical applications have shown excellent performance. As the structure of synthetic peptides becomes increasingly complex, the protective groups of amino acids tend to develop towards selectivity and specificity. Enzymatic and photolytic protective groups, with their unique advantages, may become new research hotspots in the future