鸡β-防御素-13成熟肽的原核表达与抑菌活性分析

Prokaryotic expression and antimicrobial activity analysis of chicken beta-defensin-13 mature peptide gene

为表达鸡β-防御素-13(Gal-13)成熟肽并分析其抑菌活性,本研究采用RT-PCR技术从广西黄鸡肝脏中扩增Gal-13全长基因,亚克隆其成熟肽基因并构建重组表达质粒,转化大肠杆菌BL21(DE3)进行诱导表达。结果显示,Gal-13全长为312 bp,编码60个氨基酸,经Blast比对后显示其与广西黄鸡Gal-13序列同源性高达100%。SDS-PAGE和western blot分析表明重组蛋白在大肠杆菌中得到大量表达,其分子量约为31 ku。抑菌试验检测结果显示,纯化后的重组蛋白对大肠杆菌BL21(DE3)和金黄色葡萄球菌具有一定的抑制作用。

To prokaryotically express Gal-13 mature peptide, chicken beta-defensin-13 （Gal-13） gene was amplified from liver of Guangxi Yellow chicken by RT-PCR. The sequencing result showed that the gene was 312 bp, encoding 60 amino acids. In addition, the cDNA of mature peptide was inserted into pGEX-4T-1 for expression in E.coli, and the expressed recombinant protein was identified by SDS-PAGE and western blot which showed that the molecule weight of the recombinant Gal-13 about 31 ku. Further identification indicated that the recombinant Gal-13 had antimicrobial bioactivity against E.coli and S.aureus.