Preparation method and applications of oriented immobilized PEGA composite resin

Abstract

The invention discloses a preparation method and applications of an oriented immobilized PEGA composite resin. The preparation method comprises following steps: 2-amino-6-chloropurine and 4-nitrobenzyl alcohol are taken as raw materials for Williamson ether synthesis so as to obtain 4-nitryl-O6-benzylguanine; 4-nitryl- O6-benzylguanine is subjected to reduction reaction with protection of t-butyloxycarboryl; coupling reaction of an obtained compound with gamma-aminobutyric acid protected by carbobenzoxy chloride is carried out in the presence of ethyl dimethyl carbodiimide and 1-hydroxybenzotriazole; O<6>-benzylguanine derivative modified PEGA resin is obtained via reduction, separation and purification, reaction with PEGA, and deprotection; and transalkylation reaction of the O<6>-benzylguanine derivative modified PEGA resin with a protein containing MGMT label is carried out, so that oriented immobilization on PEGA resin surface via thioether covalent bonds is realized. Reaction of the preparation method is stable; the steps are simple; a stable uniform protein coating with uniform orientation can be formed on solid material surface by a prepared product; and the preparation method is used for preparing reagent kit with specific recognition effects.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to immobilizing a recombinant protein containing O6-alkylguanine-DNA methyltransferase (MGMT) on the surface of a solid material to establish a method for immobilizing the MGMT recombinant protein. Background technique [0002] Technologies such as enzyme reactors, biosensors, and biochips centered on immobilized proteins are of great significance for biocatalysis, proteomics, clinical diagnosis, and new drug development. The strength of the immobilized protein bioactivity is the prerequisite and determinant factor for the successful construction of the above-mentioned immobilized protein interface, and it is also the most critical factor affecting the results of later catalysis, identification, diagnosis and screening. Therefore, the research on the high activity immobilization method of protein can not only improve the accuracy of the above-mentioned technical research results, but also...

AI Technical Summary

Problems solved by technology

However, due to the non-specificity of the method used and the weak interaction between the protein and the solid material, the immobilized protein obtained by this method is prone to loss during use, which also affects the accuracy of the research results

The chemical bonding method can be divided into random immobilization and directional immobilization. The random immobilization method generally uses groups such as amino groups, carboxyl groups, and sulfhydryl groups in the protein structure to chemically react with aldehyde groups modified on the surface of solid materials. The valence bond fixes the protein on the solid surface at will. Due to the different orientation and conformation of the protein on the solid surface during the fixation process, the active site of this method is easily affected.

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