介导金黄色葡萄球菌侵袭细胞的关键蛋白FnBP功能肽段在E.coli中的可溶性表达及功能

Soluble expression and preliminary study on a functional peptide of FnBP protein mediating Staphylococcus aureus invasion

目的:将介导金黄色葡萄球菌侵袭细胞的关键蛋白FnBP的重要功能肽段与GST蛋白在大肠杆菌中融合表达.方法:利用合成的特异性引物钓取金黄色葡萄球菌基因组中的目的片段,将其克隆入pGEX-KG载体,在大肠杆菌中与GST蛋白进行融合表达.结果:成功地表达了目的肽,可溶性的表达产物占细菌可溶性蛋白的20%左右,且目的肽与配基纤连蛋白Fn具有结合的能力.结论:成功地表达了介导金黄色葡萄球菌侵袭细胞的关键蛋白FnBP中的重要功能肽段,为研究金葡菌侵袭细胞的机制奠定了基础.

AIM： to express the gene fragment of interesting peptide that comprised of C-terminal D1 and N-terminal D2 （FnBP-Dlc/D2n） from the genome of 04018 in E. coli in GST- fusion form by use of DNA recombinant technique and genetic engineering. METHODS： The specific primers were synthesized to extract the fragment of interesting peptide from the genome of the staphylococcus aureus, then the gene fragment of interesting peptide was correctly cloned into expression vector pGEX-KG. The recombinant plasmid pGEX-KG was introduced into E. coli DH5α that was introduced to express the protein in GST-fusion form. RESULTS： Reconstructed gene could be expressed effectively in E. coli in soluble form. The molecular weight of expressed product of recombinantion plasmid pGEX-KG analyzed by SDS-PAGE was the same as anticipated. And the soluble expresssion product accounted for 20% of the total bacterial protein. ELISA assay showed that the expression product could bind to fibronectin. CONCLUSION： The expression vector has been constructed and reconstructed gene was expressed successfully and effectively in E. coli, which may provide a useful basis on developing new therapeutic and vaccine strategies for S. aureus invasion of cells.