HSF1中的半胱氨酸残基氧化还原状态对其功能的调节  

HSF1 may have Integrated Redox Dependent Regulation of Cysteine Residues into Its Function Response

作　　者：林正[1] 黄帆[1] 罗兰[1] 张式鸿[1] 马中富[1] 吴兴刚[1] 徐康[1] 

机构地区：[1]中山大学附属第一医院中心实验室,广州510080

出　　处：《中国生物化学与分子生物学报》2005年第3期341-346,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家自然科学基金(No.30070829);广东省自然科学基金资助项目(No.31695)~~

摘　　要：人热休克转录因子1(heatshocktranscriptionfactorl ,HSF1)的结构和功能与其半胱氨酸残基的氧化还原化学性能相关.为了鉴别在氧化还原状态改变时参与分子内双硫键交联的HSF1半胱氨酸残基,了解其氧化还原化学性改变在生物学调节中的重要性,通过建立和应用重组人HSF1中的5个半胱氨酸突变体和1个双重突变体,并用已知的巯基氧化介导剂联氨(diamide ,H2 N·NH2 )和还原介导剂二硫苏糖醇(DTT)与在体外转录和翻译的HSF1突变体蛋白质预孵育,观察其构象和与DNA结合活性的改变.结果显示,与野生型一样,所有的HSF1半胱氨酸突变体都能被热激活并与DNA结合;联氨预处理能阻断这种作用,但对突变体C153 S和双重突变体C3 73、3 78S无效.氧化还原状态对HSF1构象改变显示联氨能使HSF1野生型和突变体C3 6T和C10 3 Y形成氧化型HSF1(ox HSF1)构象,但对C153 S和C3 73、3 78S双重突变体不起作用,而单一突变体C3 73 S或C3 78S在联氨作用下分别形成二种分子量稍不同的ox HSF1构象.结果提示,在氧化条件下HSF1中的半胱氨酸残基C153 可能与C3 73 或与C3 78形成分子内二硫键交联;在对抗氧化作用上,C153 和C3 73 、C3 78起着“关闭性”作用,预防了HSF1的激活.To evaluate the fundamentally important role of redox status of human heat shock transcription factor 1 (HSF1) in biological regulation, cysteine site-specific mutants of HSF1 were constructed and used to determine which of the five cysteine residues may be engaged in disulfide cross-link. Analysis of the in vitro transcribed and translated HSF1 proteins showed that while mutation of C-~36 and C-~103 had no effect on the redox sensitivity of HSF1, the mutation of C-~153 or double mutation of C-~373 and C-~378 to serine rendered HSF1 insensitive to oxidizing reagent diamide and prevented its conversion to ox-HSF1. HSF1 with a single cysteine to serine mutation at either the C-~373 or C-~378 position obtained different ox-HSF1 conformers in the presence of diamide. The results suggest that cysteines residues of HSF1, particularly C-~153 and C-~373 ,C-~378 are necessary and sufficient to form an intramolecular disulfide cross-link under an oxidizing condition and likely provide an off-switch triggered by oxidation to prevent the activation of HSF1.

关 键 词：热休克转录因子1 半胱氨酸 突变体 氧化还原 

分 类 号：Q256[生物学—细胞生物学] Q71