| 烟草硫氧还蛋白NtTRXh功能域研究 |
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| 引用本文： | 刘会敏,孙力军,任海英,董汉松.烟草硫氧还蛋白NtTRXh功能域研究[J].江苏农业学报,2010,26(2). |
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| 作者姓名： | 刘会敏 孙力军 任海英 董汉松 |
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| 作者单位： | 南京农业大学植物保护学院,江苏,南京,210095 |
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| 基金项目： | 国家自然科学基金项目(30771441) |
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| 摘 要： | 利用PCR点突变的方法对烟草硫氧还蛋白基因(NtTRX)进行点突变,得到3个突变克隆和突变蛋白。把N端第13位半胱氨酸(Cysteine,C)残基(13-C)以及第68位、并位于WCGPC中的半胱氨酸残基(68-C)分别用苏氨酸(Threonine,T)加以替换,得到突变蛋白C13T和C68T;用T对13-C和68-C进行双突变,得到突变蛋白C13TC68T。通过原核表达体系表达产生了相应的蛋白质,并分别进行了纯化。利用硫氧还蛋白能催化胰岛素二硫键还原的性质,比较了C13T、C68T、C13TC68T和NtTRXh的活性。结果表明,只有NtTRXh显示活性,3个突变蛋白都失去了还原二硫键的能力。可见,除了WCGPC中的C外,NtTRXh氮末端的C残基也是催化反应所必需的。
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| 关 键 词： | 硫氧还蛋白 活性位点 点突变 二硫键 |
Analysis of the Functional Domain of a Thioredoxin Protein from Tobacco |
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| LIU Hui-min,SUN Li-jun,REN Hai-ying,DONG Han-song.Analysis of the Functional Domain of a Thioredoxin Protein from Tobacco[J].Jiangsu Journal of Agricultural Sciences,2010,26(2). |
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| Authors: | LIU Hui-min SUN Li-jun REN Hai-ying DONG Han-song |
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| Institution: | College of Plant Protection/a>;Nanjing Agricultural University/a>;Nanjing 210095/a>;China |
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| Abstract: | Site-directed mutation protocols were applied to the NtTRX gene to induce changes in the codon for 13-cysteine(C) located at N-terminus of the gene,in the codon for 68-C residue present in the consensus WCGPC,and in both codons.The protocols generated mutant gene clones C13T,C68T and C13TC68T,which were anticipated to encode mutant proteins in 13-C,68-C,and both were replaced with threonine(T).After the gene mutations were confirmed based on sequencing information,the three mutant proteins and the parent pr... |
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| Keywords: | thioredoxin catalytic site site-directed mutation disulfide bond |
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