关键词: 金黄色葡萄球菌, 噬菌体, 生物信息分析, 内溶素, 克隆表达

Abstract: This study aimed to obtain the endolysin lys56 encoded by Staphylococcus aureus phage qdsa001. First of all, the characteristics (physicochemical properties, signal peptide, transmembrane helices and structure domains) of lys56 were predicted by several online software. Subsequently, the target protein was obtained by cloning and expression. For this purpose, the target gene was synthesized and inserted into a prokaryotic expression vector pET-30a to construct recombinant plasmid pET-30a-lys56, which was subsequently transformed into E. coli Rosetta (DE3). Ni-IDA affinity chromatography was applied to purify the recombinant protein. The results showed that lys56 had a relative molecular mass of 35.1 kDa without signal peptide or transmembrane domain and contained only one Ami_2 domain. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed that the recombinant protein was successfully expressed in the cell supernatant with a molecular mass of about 35 kDa, which was consistent with the expected size. Optimal expression of the endolysin lys56 was induced by 0.5 mmol/L isopropyl-β-D-thiogalactopyranoside (IPTG) overnight at 20 ℃.

Key words: Staphylococcus aureus, bacteriophage, bioinformatics analysis, endolysin, cloning and expression