...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Biochemistry >Molecular dissection of the folding mechanism of the alpha subunit of tryptophan synthase: an amino-terminal autonomous folding unit controls several rate-limiting steps in the folding of a single domain protein.
【24h】

Molecular dissection of the folding mechanism of the alpha subunit of tryptophan synthase: an amino-terminal autonomous folding unit controls several rate-limiting steps in the folding of a single domain protein.

机译：色氨酸合酶的α亚基折叠机制的分子解剖：氨基末端自主折叠单元控制单个结构域蛋白折叠中的几个限速步骤。

获取原文
获取原文并翻译 | 示例
   

获取外文期刊封面封底 >>

       
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

The alpha subunit of tryptophan synthase (alphaTS) from Escherichia coli is a 268-residue 8-stranded beta/alpha barrel protein. Two autonomous folding units, comprising the first six strands (residues 1-188) and the last two strands (residues 189-268), have been previously identified in this single structural domain protein by tryptic digestion [Higgins, W., Fairwell, T., and Miles, E. W. (1979) Biochemistry 18, 4827-4835]. The larger, amino-terminal fragment, alphaTS(1-188), was overexpressed and independently purified, and its equilibrium and kinetic folding properties were studied by absorbance, fluorescence, and near- and far-UV circular dichroism spectroscopies. The native state of the fragment unfolds cooperatively in an apparent two-state transition with a stability of 3.98 +/- 0.19 kcal mol(-1) in the absence of denaturant and a corresponding m value of 1.07 +/- 0.05 kcal mol(-1) M(-1). Similar to the full-length protein, the unfolding of the fragment shows two kinetic phases which arise from the presence of two discrete native state populations. Additionally, the fragment exhibits a significant burst phase in unfolding, indicating that a fraction of the folded state ensemble under native conditions has properties similar to those of the equilibrium intermediate populated at 3 M urea in full-length alphaTS. Refolding of alphaTS(1-188) is also complex, exhibiting two detectable kinetic phases and a burst phase that is complete within 5 ms. The two slowest isomerization phases observed in the refolding of the full-length protein are absent in the fragment, suggesting that these phases reflect contributions from the carboxy-terminal segment. The folding mechanism of alphaTS(1-188) appears to be a simplified version of the mechanism for the full-length protein [Bilsel, O., Zitzewitz, J. A., Bowers, K.E, and Matthews, C. R.(1999) Biochemistry 38, 1018-1029]. Four parallel channels in the full-length protein are reduced to a pair of channels that most likely reflect a cis/trans proline isomerization reaction in the amino-terminal fragment. The off- and on-pathway intermediates that exist for both full-length alphaTS and alphaTS(1-188) may reflect the preponderance of local interactions in the beta/alpha barrel motif.
机译：来自大肠杆菌的色氨酸合酶（alphaTS）的alpha亚基是268个残基的8链beta / alpha桶状蛋白。先前已通过胰蛋白酶消化在该单个结构域蛋白中鉴定了两个自主折叠单元，包括前六链（残基1-188）和后两链（残基189-268）[Higgins，W.，Fairwell，T ，和Miles，EW（1979）Biochemistry 18，4827-4835]。较大的氨基末端片段alphaTS（1-188）被过表达并独立纯化，并且通过吸光度，荧光以及近和远紫外圆二色性光谱研究了其平衡和动力学折叠特性。片段的原始状态在明显的两态过渡中协同展开，在没有变性剂的情况下稳定性为3.98 +/- 0.19 kcal mol（-1），相应的m值为1.07 +/- 0.05 kcal mol（- 1）M（-1）。与全长蛋白质相似，片段的展开显示出两个动力学相，这是由于存在两个离散的天然状态种群而引起的。另外，该片段在展开中表现出显着的猝发相，表明在自然条件下一部分折叠状态集合具有类似于全长αTS中填充于3M尿素的平衡中间体的那些性质。 alphaTS（1-188）的重折叠也很复杂，表现出两个可检测的动力学阶段和一个在5毫秒内完成的猝发阶段。在全长蛋白质的重折叠中观察到的两个最慢的异构化阶段不存在于片段中，表明这些阶段反映了羧基末端片段的贡献。 alphaTS（1-188）的折叠机制似乎是全长蛋白质机制的简化版本[Bilsel，O.，Zitzewitz，JA，Bowers，KE，and Matthews，CR（1999）Biochemistry 38，1018 -1029]。全长蛋白质中的四个平行通道被还原为一对通道，最有可能反映出氨基末端片段中的顺式/反式脯氨酸异构化反应。全长alphaTS和alphaTS（1-188）都存在的非通路中间体和通路中间体可能反映了beta / alpha桶基序中局部相互作用的优势。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号