[Redox-dependent changes in structure and function of hHSF1].

Zheng Lin, Fan Huang, Zhong-Fu Ma, Kang Xu, Alice Y Liu
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Abstract

To evaluate the effects of cysteine-SH-directed regents on the redox status, structure and function of human heat shock transcription factor 1 (hHSF1), treatment in vitro of hHSF1 with 0.3 0.5 mmol/L oxidizing reagent diamide (DM) and treatment in vivo of HeLa cells with 1 mmol/L buthionine sulfoximine (BSO), an inhibitor of gamma-glutamylcysteine synthetase, promoted the formation of a compact, intramolecularly disulfide-crosslinked, stable monomeric form of ox-hHSF1, and blocked the trimerization and activation of HSF1. The effects of diamine were dose-dependent and readily could be completely reversed by adding 0.4 0.5 mmol/L reducing reagent dithiothreitol (DTT) to the samples prior to gel electrophoresis. Computer modeling of the alpha-helical coiled-coil domains of the HSF1 monomer and trimer showed that the alignment of the N- and C-terminal hydrophobic repeats of HSF1 monomer could bring C(3)(Cys(153))close to C(4) and C(5)(Cys(373) and Cys(378), respectively), in positions permissible for disulfide bond formation under appropriate experimental conditions. The results suggest that redox-dependent thiol-disulfide exchange can provide a mechanism for regulation the conformation and activity of hHSF1.

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[氧化还原依赖性hHSF1结构和功能的变化]。
为了评价半胱氨酸- sh定向试剂对人热休克转录因子1 (hHSF1)氧化还原状态、结构和功能的影响,在体外用0.3 - 0.5 mmol/L氧化试剂二胺(DM)处理hHSF1,在体内用1 mmol/L γ -谷氨酰基半胱氨酸合成酶抑制剂丁硫氨酸亚砜(BSO)处理HeLa细胞,促进了ox-hHSF1分子内紧密、二硫交联、稳定的单体形式的形成。阻断HSF1的三聚体化和活化。凝胶电泳前加入0.4 ~ 0.5 mmol/L还原试剂二硫苏糖醇(DTT)即可完全逆转二胺的作用。对HSF1单体和三聚体的α -螺旋螺旋结构域的计算机模拟表明,HSF1单体的N端和C端疏水重复序列的排列可以使C(3)(Cys(153))接近C(4)和C(5)(Cys(373)和Cys(378)),在适当的实验条件下允许形成二硫键的位置。结果表明,氧化还原依赖的硫醇-二硫交换可能为调节hHSF1的构象和活性提供了一种机制。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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