How to choose peptide synthesis process?
Chemical synthesis of peptides is a very special branch of organic synthesis. At present, there are two main methods: liquid phase synthesis and solid phase synthesis.
Liquid phase synthesis is a classical peptide synthesis method, which generally adopts step-by-step synthesis or fragment condensation. The step-by-step synthesis method usually starts from the C 'terminal amino acid of the chain and repeatedly adds a single amino acid to the increasing amino acid component α- Amino protected amino acids. Fragment condensation generally divides the target sequence into fragments reasonably, then synthesizes each fragment step by step, and finally condenses each fragment according to the sequence requirements. The advantages of liquid phase synthesis are that the intermediate products can be purified in each step, the physicochemical constants of the intermediate products can be obtained, non amino acid modification can be carried out at will, and amino acid deletion can be avoided. The disadvantages are time-consuming and laborious.
Solid phase synthesis is to connect the carboxyl group of the first amino acid of the target peptide with the solid-phase carrier (resin) in the form of covalent bond, and then take the amino group of this amino acid as the synthesis starting point to acylate it with the carboxyl group of adjacent amino acids (amino protection) to form peptide bond. Then, the amino group of the resin peptide containing these two amino acids is deprotected and reacts with the carboxyl group of the next amino acid, and this process is repeated until the target peptide is formed. The advantages are that the post-treatment operation of each reaction step is simplified, the loss caused by manual operation and material transfer is avoided, the yield is high, and automation can be realized; The disadvantage is that the intermediate product in each step can not be purified, large amino acid excess feeding must be adopted, the purity of crude product is not as high as that of liquid phase compound, and it must be purified by reliable separation means.
Liquid phase synthesis and solid-phase synthesis have their own advantages and disadvantages. The appropriate process should be selected according to the actual needs of synthesis. Generally speaking, liquid phase synthesis is more suitable for the synthesis of short peptides; Solid phase synthesis is more suitable for the synthesis of medium and long peptides. Of course, the two methods can also be combined. For example, the short peptide fragment can be synthesized by liquid-phase method, and then the fragment can be applied to solid-phase synthesis.
Whether liquid phase synthesis or solid phase synthesis, the target molecule is obtained by directional formation of amide bond according to the designed amino acid sequence. Theoretically speaking, this is not complicated, but there are still many factors to be considered in implementation. A simple carboxylic acid forms an amide bond with an amine. Generally, the carboxyl group is first transformed into an active carboxyl derivative (such as acyl chloride or anhydride) and then reacts with the amine, or a condensation agent is added to the reaction system. However, the formation of amide bonds between amino acids is much more complex because each amino acid contains both amino and carboxyl groups. If the carboxyl group of an amino acid is activated, it can react with the amino group of the same or another amino acid molecule; If several amino acids are mixed together and a condensation agent is added, only a mixture composed of polypeptides with a variety of different amino acid sequences can be obtained. Therefore, in the research of peptide synthesis, we should not only pay attention to activation methods and coupling methods, but also pay attention to the selection of protection / deprotection strategies.
In the process of peptide synthesis, some heteropeptides with similar structure to the target peptide will be produced, such as diastereomers due to amino acid racemization, missing peptides due to the lack of connection of some amino acids, broken peptides due to peptide bond breakage, etc. Therefore, it is necessary to consider selecting reliable separation and purification methods to make the purity of peptides meet the requirements. The purification of polypeptide drugs usually adopts chromatography, but in some cases, the common purification methods in organic synthesis (such as recrystallization) may also be applicable.
