变铅青链霉菌广谱胁迫蛋白对氧化压力响应的研究

Response of universal stress protein from Streptomyces lividans 1326 to oxidative stress

摘要:

【目的】广谱胁迫蛋白(USP)是一种古老的蛋白家族，在链霉菌属细菌中其功能研究尚未报道。以变铅青链霉菌USP蛋白为对象对其功能进行解析。【方法】使用序列比对的方法分析同源性及保守结构域。纯化USP蛋白，用圆二色谱分析蛋白与环腺苷酸(cAMP)的结合对usp (SLI_7517)进行基因中断。检测野生型和usp基因缺失株对偶氮二甲酰胺造成的氧化压力的耐受能力。使用qPCR荧光定量分析技术，检测野生型菌株与usp缺失株在氧化环境中谷胱甘肽过氧化物酶及巯基过氧化物酶基因转录量的差异。【结果】同源序列分析表明链霉菌属来源的USP蛋白序列相互之间相似性较高，USP-like结构域高度保守。USP蛋白在体外结合cAMP引起CD谱的变化。usp基因缺失株对偶氮二甲酰胺更耐受，同时菌株中谷胱甘肽过氧化物酶基因转录量上升。【结论】变铅青链霉菌中USP蛋白能够结合cAMP。usp参与菌体应对氧化环境的调控，对谷胱甘肽过氧化物酶基因的转录有阻遏作用。

Abstract:

[Objective] Universal stress protein (USP) belongs to an ancient protein family. In Streptomyces, its physiological function has not been reported yet. This work focused on the protein particularly. [Methods] Sequence alignment was performed. The association between cAMP and USP was detected using circular dichroism spectroscopy. The usp gene (SLI_7517) was also disrupted. Susceptibility of wild-type and the usp disruption mutant strains to the thiol oxidant, diamide, was analyzed. qPCR was conducted to detect the expression differences of glutathione peroxidase and thiol peroxidase genes between the wild-type and usp disruption mutant strains. [Results] USP from different Streptomyces species shares 90% similarity, with the highly conserved USP-like domain. The alteration of USP CD spectra was observed when USP was mixed with cAMP. Tolerance to diamide increased after usp was knocked out. The transcription level of glutathione peroxidase gene was up-regulated in the usp disruption mutant. [Conclusion] USP protein from Streptomyces lividans 1326 can bind to cAMP. It might be involved in the regulation process, that suppresses the transcription of glutathione peroxidase when the bacteria exposed to oxidative environment.