Peptide synthesis knowledge grocery store side chain protection group, synthesis detection

Peptide chemistry is the flexible use and collocation of amino acid protective groups

For the use of side chain protecting groups, please refer to Chapter 4 of “solid phase organic synthesis – Principles and Application guide” edited by Wang Dexin. This paper mainly introduces several protective groups of Cys, Lys and ASP and their removal methods.

The most common protective groups of Cys are TRT, ACM and mob, which can complete the synthesis of multiple disulfide bond peptides. The most common protective groups of Lys are BOC, Fmoc, TRT, DDE, allyl, which provide many orthogonal protection strategies for solid-phase synthesis of cyclic peptides. The most common protective groups of ASP are otbu, Obzl, ome, oall and OFM, which also provide a variety of orthogonal protection strategies.

Common protective groups of sulfhydryl

Ninhydrin detection

In solid-phase peptide synthesis, the connection efficiency is mainly determined by detecting the free amino groups on the resin. Kaiser was used for biological detection.

The detection results, if there is free amino group, show blue, or reddish brown (pro, Ser, his). Kaiser reagent includes: A, 6% ninhydrin ethanol solution B, 80% phenol ethanol solution C, 2% 0.001M KCN pyridine solution. The pyridine in the preparation needs to be treated with ninhydrin and then re evaporated before use.

In the detection process, take a small amount of resin, add 2 drops of a, B and C, and heat it at 110 ℃ for 3 min. if the solution has blue color, or the resin appears blue or reddish brown, it indicates that there are free amino groups, otherwise, it indicates that the connection is complete. There are other methods to detect free amino groups: bromofinland method, etc.

Detection principle of ninhydrin

Cutting method of solid phase synthesis

After the solid-phase peptide synthesis is completed, it is necessary to select the appropriate cutting reagent to cut the peptide from the resin, and then through ether precipitation, centrifugation and precipitation, separation and purification by HPLC, and freeze-drying to obtain the final peptide product.

Antonius bio reminds that due to the different resin and amino acid sequence, the cutting method is not the same. Generally, the acid conditions are selected for cutting.

For PAM and mbha resin, HF cutting is generally used, and reagents need to be added in the cutting process. For Wang, rink amide and TRT resins, TFA is generally used for cutting. In the process of cutting, ethylene dithiol, phenyl sulfate and water are added.

These additives are mainly used as carbon positive ion capture reagents, in order to capture the carbon positive ions generated in the cutting reaction process, and reduce the side reactions caused by the attack of these carbon positive ions on the side chain of some amino acids.

The amino acids which are easy to produce side effects are Trp and Tyr. The amount of cutting reagent is generally 10-15ml / g resin. The commonly used cutting ratio is HF / p-cresol / P-thiocresol (90 / 5 / 5), TFA / tis / EDT / H2O (94 / 1 / 2.5 / 2.5), and the reaction is generally 2 H-4 h at room temperature.

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