...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Biochemistry >A Single Mutation Traps a Half-Sites Reactive Enzyme in Midstream, Explaining Asymmetry in Hydride Transfer
【24h】

A Single Mutation Traps a Half-Sites Reactive Enzyme in Midstream, Explaining Asymmetry in Hydride Transfer

机译：单一突变捕获半场位点在中间的反应性酶，解释氢化物转移中的不对称性

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

In Escherichia coli thymidylate synthase (EcTS), rate-determining hydride transfer from the cofactor 5,10-methylene-5,6,7,8-tetrahydrofolate to the intermediate 5-methylene-2′-deoxyuridine 5′-monophosphate occurs by hydrogen tunneling, requiring precise alignment of reactants and a closed binding cavity, sealed by the C-terminal carboxyl group. Mutations that destabilize the closed conformation of the binding cavity allow small molecules such as β-mercaptoethanol (β-ME) to enter the active site and compete with hydride for addition to the 5-methylene group of the intermediate. The C-terminal deletion mutant of EcTS produced the β-ME adduct in proportions that varied dramatically with cofactor concentration, from 50% at low cofactor concentrations to 0% at saturating cofactor conditions, suggesting communication between active sites. We report the 2.4 ? X-ray structure of the C-terminal deletion mutant of E. coli TS in complex with a substrate and a cofactor analogue, CB3717. The structure is asymmetric, with reactants aligned in a manner consistent with hydride transfer in only one active site. In the second site, CB3717 has shifted to a site where the normal cofactor would be unlikely to form 5-methylene-2′-deoxyuridine 5′-monophosphate, consistent with no formation of the β-ME adduct. The structure shows how the binding of the cofactor at one site triggers hydride transfer and borrows needed stabilization from substrate binding at the second site. It indicates pathways through the dimer interface that contribute to allostery relevant to half-sites reactivity.
机译：在大肠杆菌胸苷替代合成酶（ECTS）中，将氢化物从辅因子5,10-亚甲基-5,6,7,8-四氢醇转移到中间体5-亚甲基-2'-脱氧酸5'-单磷酸盐中的氢化物转移通过氢隧道发生，需要精确对准反应物和封闭的结合腔，由C末端羧基密封。破坏结合腔的闭合构象的突变允许小分子如β-巯基乙醇（β-ME）进入活性位点并与氢化物竞争，以除去中间体的5-亚甲基。 ECTS的C末端缺失突变体以比例产生的β-ME加合物，其比例随着辅因子浓度而变化，在饱和辅因子条件下在低辅因子浓度下为0％，表明活性位点之间的通信。我们报告了2.4？ e的C末端缺失突变体的X射线结构。 Coli Ts与底物和辅助弧剂类似物，CB3717复合物。该结构是不对称的，反应物以一致的方式对准仅在一个活性位点中的氢化物转移。在第二站点中，CB3717已经转移到正常辅因子不太可能形成5-亚甲基-2'-脱氧尿苷5'-一磷酸的部位，这与β-ME加合物的不形成一致。该结构表明了辅因子在一个部位的结合如何触发氢化物转移和胆汁从第二位点处的基材结合稳定稳定。它表明了通过二聚体界面的途径，这些界面有助于与半场反应性相关的仿生。

著录项

  • 来源
    《Biochemistry》 |2018年第19期|共10页
  • 作者

    Janet S. Finer-Moore; Tom T. Lee; Robert M. Stroud;

  • 作者单位

    Department of Biochemistry and Biophysics University of California San Francisco California 94143-2240 United States;

    Department of Biochemistry and Biophysics University of California San Francisco California 94143-2240 United States;

    Department of Biochemistry and Biophysics University of California San Francisco California 94143-2240 United States;

  • 收录信息
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类 生物化学;
  • 关键词

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