...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Biochemistry >Alteration of the Flexible Loop in 1?Deoxy?D?xylulose-5-phosphate Reductoisomerase Boosts Enthalpy-Driven Inhibition by Fosmidomycin
【24h】

Alteration of the Flexible Loop in 1?Deoxy?D?xylulose-5-phosphate Reductoisomerase Boosts Enthalpy-Driven Inhibition by Fosmidomycin

机译：1？脱氧？D？木糖基5-磷酸还原异构酶中柔性环的改变增强了膦霉素对焓驱动的抑制作用

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

1-Deoxy-D-xylulose-5-phosphate reductoisomerase (DXR), which catalyzes the first committed step in the 2-C-methyl-D-erythritol 4-phosphate pathway of isoprenoid biosynthesis used by Mycobacterium tuberculosis and other infectious microorganisms, is absent in humans and therefore an attractive drug target. Fosmidomycin is a nanomolar inhibitor of DXR, but despite great efforts, few analogues with comparable potency have been developed. DXR contains a strictly conserved residue, Trp203, within a flexible loop that closes over and interacts with the bound inhibitor. We report that while mutation to Ala or Gly abolishes activity, mutation to Phe and Tyr only modestly impacts kcat and Km. Moreover, presteady- state kinetics and primary deuterium kinetic isotope effects indicate that while turnover is largely limited by product release for the wild-type enzyme, chemistry is significantly more rate-limiting for W203F and W203Y. Surprisingly, these mutants are more sensitive to inhibition by fosmidomycin, resulting in Km/Ki ratios up to 19-fold higher than that of wild-type DXR. In agreement, isothermal titration calorimetry revealed that fosmidomycin binds up to 11-fold more tightly to these mutants. Most strikingly, mutation strongly tips the entropy?enthalpy balance of total binding energy from 50% to 75% and 91% enthalpy in W203F and W203Y, respectively. X-ray crystal structures suggest that these enthalpy differences may be linked to differences in hydrogen bond interactions involving a water network connecting fosmidomycin’s phosphonate group to the protein. These results confirm the importance of the flexible loop, in particular Trp203, in ligand binding and suggest that improved inhibitor affinity may be obtained against the wild-type protein by introducing interactions with this loop and/or the surrounding structured water network.
机译：1-脱氧-D-木酮糖-5-磷酸还原异构酶（DXR）催化结核分枝杆菌和其他传染性微生物使用的类异戊二烯生物合成的2-C-甲基-D-赤藓糖醇4-磷酸途径中的第一步。在人类中不存在，因此是有吸引力的药物靶标。磷霉素是DXR的纳摩尔抑制剂，但尽管付出了巨大的努力，但几乎没有开发出具有类似效力的类似物。 DXR在一个柔性环中包含一个严格保守的残基Trp203，该环封闭并与结合的抑制剂相互作用。我们报告说，虽然突变为Ala或Gly取消了活性，但突变为Phe和Tyr仅适度地影响了kcat和Km。此外，稳态前动力学和氘代动力学同位素效应表明，虽然营业额在很大程度上受到野生型酶产物释放的限制，但化学性质对W203F和W203Y的限速性明显更大。令人惊讶的是，这些突变体对磷霉素的抑制作用更加敏感，导致Km / Ki比野生型DXR高19倍。一致的是，等温滴定量热法显示，磷霉素与这些突变体的结合多达11倍。最显着的是，突变强烈提示W203F和W203Y的总结合能的熵焓焓分别从50％增至75％和91％。 X射线晶体结构表明，这些焓差可能与氢键相互作用的差异有关，氢键相互作用包括将膦酰胺膦酸酯基团与蛋白质连接的水网络。这些结果证实了柔性环，特别是Trp203，在配体结合中的重要性，并表明通过引入与该环和/或周围结构化水网络的相互作用，可以获得对野生型蛋白的改善的抑制剂亲和力。

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