鼠伤寒沙门菌毒力基因SptP的原核表达及生物信息学分析

doi:10.16431/j.cnki.1671-7236.2022.12.023

Abstract

Abstract: 【Objective】 The purpose of the experiment was to obtain the highly expressed SptP protein of Salmonella Typhimurium and carry out bioinformatics analysis, so as to provide a theoretical basis for its function research and screening of interacting proteins.【Method】 The SptP gene was amplified by PCR technology, and the sequence was ligated to pET-32a(+) vector to construct the prokaryotic expression vector pET32a-SptP of Salmonella Typhimurium SptP gene.The recombinant plasmid was introduced into Escherichia coli BL21(DE3) competent cells by heat shock method, the recombinant protein was induced to express by IPTG, purified, and verified by SDS-PAGE and Western blotting.Then the SptP protein was analyzed by bioinformatics using online softwares.【Result】 The SptP gene was successfully amplified by PCR with a size of 1 632 bp.The recombinant SptP protein was successfully induced, expressed and purified in E.coli BL21(DE3) competent cells to obtain a protein with a molecular mass of 79.7 ku.The molecular formula of the SptP protein was C₂₆₂₅H₄₂₅₇N₇₄₅O₈₁₂S₂₅, no transmembrane structure, no signal peptide, and a molecular mass of 60 047.68 u, the theoretical isoelectric point was 8.75, and there were 57 possible phosphorylation sites.The SptP protein was mainly located in the nucleus, cytoplasm, Golgi apparatus, cytoskeleton, secretion system vesicles, and plasma membrane, accounting for 43.5%, 34.8%, 8.7%, 4.3%, 4.3% and 4.3%, respectively.SptP protein was mainly composed of alpha helix, extended strand, beta turn and random coil, accounting for 43.65%, 14.92%, 4.42% and 37.02%, respectively.【Conclusion】 In this study, a recombinant plasmid pET32a-SptP expressing SptP protein was constructed, and a SptP recombinant protein with a molecular mass of 79.7 ku was obtained.The basic physical and chemical properties and biological functions of SptP protein were clarified, which provided theoretical and experimental basis for the interaction mechanism between SptP protein and host cells and the preparation of new Salmonella Typhimurium vaccine.

Key words: Salmonella Typhimurium; SptP; prokaryotic expression; bioinformatics analysis

CLC Number:

S852.61^＋2

ZHOU Nanlong, DING Yonghui, LI Tiansen. Prokaryotic Expression and Bioinformatics Analysis of Salmonella Typhimurium Virulence Gene SptP[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(12): 4734-4744.

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Prokaryotic Expression and Bioinformatics Analysis of Salmonella Typhimurium Virulence Gene SptP

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