融合超嗜热菌Pyrococcus furiosus红素氧还蛋白可提高靶蛋白的热稳定性  

Improvement on the Thermostability of Target Proteins by Fusing Rubredoxin from Hyperthermophile Pyrococcus furiosus

作　　者：吴娇 余桂珍 袁航[1] 刘娴[1] 高艳秀[1] 龚明[1] 邹竹荣[1] WU Jiao;YU Gui-zhen;YUAN Hang;LIU Xian;GAO Yan-xiu;GONG Ming;ZOU Zhu-rong(Yunnan Key Laboratory of Biomass Energy and Environmental Biotechnology,Ministry of Education,Research Center for Sustainable Development and Applied Engineering of Biomass Energy,School of Life Sciences,Yunnan Normal University,Kunming 650500)

机构地区：[1]云南师范大学生命科学学院云南省生物质能源和环境生物技术重点实验室教育部生物质能源持续发展和应用工程研究中心,昆明650500

出　　处：《生物技术通报》2021年第10期110-119,共10页Biotechnology Bulletin

基　　金：国家自然科学基金项目(31760077)。

摘　　要：热稳定性是功能蛋白(特别是酶蛋白)储存和使用的重要限制因子。许多商业酶因耐热性不能达到工业生产的热处理条件而无法被采用;另外,植物中与光合作用有关的关键酶也受高温抑制,从而导致农作物产量下降。因此,对酶蛋白进行分子改良以提高其热稳定性,进而提升其功效和扩大其应用范围,具有重要的现实意义。选取3种典型的热稳定性差、易聚集但具有重要功能的酶蛋白(小桐子抗坏血酸过氧化物酶1即JcAPX1、大肠杆菌高丝氨酸-O-转琥珀酰酶即EcMetA、假单胞菌亚磷酸盐脱氢酶即PsPtxD)作为靶蛋白,鉴定出超嗜热菌Pyrococcus furiosus的小分子红素氧还蛋白(rubredoxin)能够作为热稳定融合标签,提高大肠杆菌重组表达靶蛋白的可溶性、热稳定性及其活性的耐热能力,为通过融合表达策略提高目标蛋白的热稳定性并强化其应用提供了一个新的技术选择。Thermostability is an important restriction factor for the storage and use of functional proteins(especially enzymatic proteins).Many commercial enzymes are not suitable for use because their heat tolerance usually cannot meet the heat processing conditions of industrial production.In addition,some key enzymes involved in plant photosynthesis are also inhibited by high temperature,thus leading to the decrease of crop yield.Therefore,it is of great practical significance to improve the thermostability of enzyme proteins by molecular modification so as to enhance their efficacy and expand their application.Herein,we chose three important enzymes that are canonically heat-labile or prone to aggregation,including JcAPX1(cytosolic ascorbate peroxidase 1 of Jatropha curcas),EcMetA(homoserine-O-transsuccinidase of Escherichia coli),and PsPtxD(phosphite dehydrogenase of Pseudomonas sp.),and used them as target proteins and identified the small rubredoxin from hyperthermophile Pyrococcus furiosus as thermo-stable fusion tag.It could be used to improve the solubility,thermostability and active heat-tolerance of those recombinant target proteins expressed in E.coli.Prospectively,this might provide a new technique choice for improving the thermostability and related applications of target proteins through the fusion expression strategy.

关 键 词：蛋白热稳定性 融合标签 超嗜热菌 PYROCOCCUS furiosus 红素氧还蛋白 

分 类 号：Q936[生物学—微生物学]