酿酒酵母表达侧耳源单基因生物合成麦角硫因

Expression in Saccharomyces cerevisiae of Single Ergothioneine Synthase Genes from Pleurotus

运用生物信息学方法对侧耳属蘑菇(平菇、杏鲍菇和白灵菇)的麦角硫因合成酶基因Egt 1进行挖掘,对其蛋白结构和功能进行预测,并构建酿酒酵母表达载体,在酿酒酵母EC 1118中进行表达研究。结果表明:克隆得到侧耳属平菇、杏鲍菇和白灵菇麦角硫因合成基因PoEgt 1、PeEgt 1和PtEgt 1,三者核苷酸序列同源性为97.03%,氨基酸序列同源性为97.93%,联合菌体破碎法并通过体外酶促反应后检测到麦角硫因,产量为(2.5±0.08)mg/L,证明了该基因具有单基因合成麦角硫因的活性。

In this study,the ergothioneine synthase gene Egt 1 of Pleurotus was excavated and the structure and function of its encoded protein were predicted by using bioinformatics methods.Then,an expression vector of Saccharomyces cerevisiae was constructed,and functional verification in S.cerevisiae EC 1118 was performed.The ergothioneine synthase genes PoEgt 1,PeEgt 1 and PtEgt 1 from P.ostreatus,P.eryngii and P.tuoliensis were cloned.Among these genes,the homology of nucleotide sequences was 97.03%and the homology of amino acid sequences was 97.93%.As determined by cell disruption and in vitro enzymatic reaction,the yield of ergothioneine was(2.5±0.08)mg/L,which proved that the Egt 1 gene had the activity to synthesize ergothioneine.