贻贝足蛋白fp-5在大肠杆菌中的表达、修饰与功能分析  

Expression,modification and functional analysis of mussel foot protein fp-5in Escherichia coli

作　　者：姚林 谢紫莎 王瑞 张鲁嘉[3] 李莎 YAO Lin;XIE Zisha;WANG Rui;ZHANG Lujia;LI Sha(College of Food Science and Light Industry,Nanjing Tech University,Nanjing 211800,China;State Key Laboratory of Materials-Oriented Chemical Engineering,Nanjing Tech University,Nanjing 211800,China;School of Chemistry and Molecular Engineering,East China Normal University,Shanghai 201100,China)

机构地区：[1]南京工业大学食品与轻工学院,江苏南京211800 [2]南京工业大学材料化学工程国家重点实验室,江苏南京211800 [3]华东师范大学化学与分子工程学院,上海201100

出　　处：《生物加工过程》2024年第1期25-32,共8页Chinese Journal of Bioprocess Engineering

基　　金：国家重点研发计划(2019YFA0905203);国家自然科学基金(22178177);江苏省先进生物制造创新中心项目自主课题(XTD2201)。

摘　　要：将地中海贻贝足蛋白fp-5与不同细菌来源的酪氨酸酶在大肠杆菌中共表达,通过体内修饰获得性能改善的贻贝足蛋白。在对序列分析和密码子优化的基础上,分别构建包含酪氨酸酶与贻贝足蛋白基因的单载体和双载体共表达系统,以实现其在大肠杆菌中表达。纯化后的蛋白通过NBT/Gly染色试验和酸-硼酸盐差异光谱法分析,表明fp-5发生了不同程度的羟基化修饰,其中fp-5与来源于多刺疣微菌(Verrucomicrobium spinosum)的酪氨酸酶(TyrVs)共表达得到的5-Vs质量浓度为18.6 mg/L,修饰率达到55.20%,黏附力约为未修饰fp-5的4.6倍。与体外修饰贻贝足蛋白相比,TyrVs体内修饰得到的5-Vs具有广泛的羟基化水平以及优秀的附着力和黏附力,为生物医药等领域提供了一种有潜力的生物胶材料。The co-expression of Mytilus galloprovincialis foot protein fp-5 and tyrosinase from different bacterial sources in E.coli could produce the modified mussel foot protein with improved in vivo performance.Through sequence analysis and codon optimization,both single-vector and dual-vector co-expression systems were constructed with tyrosinase and mussel foot protein genes,and the subsequent expression proved to be successful in E.coli.The purified protein was analyzed by NBT/Gly staining test and acid-borate differential spectroscopy,showing a wide range of hydroxylation levels in the resultant fp-5.The co-expression of fp-5 and tyrosinase derived from Verrucomicrobium spinosum(TyrVs)reached a concentration of 18.6 mg/L,along with 55.2%modification rate.Moreover,the 5-Vs from such co-expression showed much higher adhesion force(4.6 times)than the unmodified fp-5.Due to the excellent adsorption and adhesion,the 5-Vs modified in vivo by TyrVs will provide a potential bio-glue material for biomedicine and other fields.

关 键 词：贻贝足蛋白 酪氨酸酶 共表达 大肠杆菌 体内修饰 

分 类 号：Q812[生物学—生物工程]