Irreversible affinity labeling agent for labeling GST and application and irreversible affinity labeling agent

Abstract

An irreversible affinity labeling agent for labeling glutathione-S-transsulfurase sj(GST) comprises an N-(dimethylhydroquinone yloxyethyl)-(N-p-substituted amido benzoyl) glycine monovalent labeling agent, and a bisamide bivalence labeling agent which is derived by the N-(dimethylhydroquinone yloxyethyl)-(N-p-substituted amido benzoyl) glycine monovalent labeling agent and dialkyl triamine; the irreversible labeling agent includes four types of functional groups, namely a dimethylhydroquinone yloxyethyl hydrophobic orienting group for guiding the labeling agent to enter a hydrophobic site at a GST active center, a modifying functional base which is represented by alpha-halogenated acyl and reacts with sulfydryl/ amino close to the GST active center, an N-bis-substituted amino acid coupling functional base which is in amide coupling with micro-nano materials, and para aminobenzoic acid or glutamic acid or acylates of para aminobenzoic acid and glutamic acid, wherein the acylates of para aminobenzoic acid and glutamic acid can supply flexible arms required by the modifying functional group for a connecting skeleton. According to the affinity labeling agent, carboxyl/ amino of the coupling functional group is in amide coupling with the micro-nano material amino/ carboxyl, and then affinity labeling is performed on sjGST, thus achieving covalent linkage outside the active bit of fusion expression protein, and as a result, a novel protein functional micro-nano material can be prepared.

Description

technical field [0001] The invention belongs to the field of protein biotechnology, and relates to an irreversible affinity labeling reagent for the active site of fusion expression tag glutathione-S-sulfurase (siGST), and coupling the affinity labeling reagent to the surface of micro-nano materials Finally, the application of site-selective immobilization of GST fusion protein. The design of this GST affinity labeling reagent is to design a bulky hydrophobic ligand for the H site of the GST active center and add a highly reactive group at an appropriate position, so that it can bind to the GST active site and be close to the active site. Specific functional groups form covalent bonds to achieve irreversible affinity labeling; coupling this affinity labeling reagent to the surface of micro-nano materials and retaining its affinity labeling activity can achieve site selectivity outside the active site of the GST fusion expression protein Immobilization to prepare protein-funct...

AI Technical Summary

Problems solved by technology

However, it is worth noting that GST is a dimer with two bulky hydrophobic binding sites, and has low selectivity for ligands; the immobilization method based on sjGST has not completely blocked its

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