Chemistry and Biology of Bacterial Sulfonucleotide Reductases

摘要 含硫氨基酸，L-半胱氨酸，是病原体的毒力和生存所必需的。生物合成用硫磺 这篇论文主要来自微生物对无机硫酸盐的同化作用。硫酸盐所需的基因- 相似对于病原体的生存是必不可少的，但在人类基因组中却不存在，因此， 治疗干预的新目标。在上一个融资周期中，我们定义了催化周期的关键特征 磺核苷酸还原酶（SR），催化硫酸盐还原的第一个关键步骤， 半胱氨酸和其它含还原硫的生物分子的合成。我们的研究提供了基本的见解 硫氧还蛋白（Trxs）--维持蛋白质硫醇还原状态的中心抗氧化酶-- 识别它们的细胞目标。我们了解到，APS还原酶（APSR）中的铁硫簇起着重要的作用， 活性位预组织和底物活化的作用，扩大了对Fe-S前体催化活性的认识， teins和PAPS还原酶（PAPR）的分化进化，缺乏这种辅因子。这些见解导致了 通过与铁硫交换中心的相互作用，以新的方式靶向APSR的分子。我们也首先发现- 硫酸盐还原的同类抑制剂，其对耐药的临床分离株表现出有效的杀菌活性， M.结核鉴于SR在病原体抗氧化应激和毒力方面的重要性，我们建议 在这里解决最大的开放问题和最大的未满足的需求仍然存在的研究领域： 化学探针，急性抑制微生物还原硫酸盐同化的基本步骤（目标1）;定义 致病性Trx促进硫还原和科普氧化还原应激的非冗余功能（目的2）;以及 在微生物半胱氨酸生物合成中的活性硫物质（RSS）的运输和输送方面的前沿知识。 总的来说，这个更新应用程序中的实验将提供有关最重要的一个方面的基本信息。 研究不足的代谢化学-还原硫酸盐同化-在一个最复杂的背景下， 病原菌M.结核我们已经建立了出色的合作关系，并在以下方面取得了良好的业绩记录： 提高生产力，以支持这些努力并确保及时完成。我们的努力将解决这些领域- 研究硫代谢在病原体抗氧化胁迫和毒力中的作用， 尽管它们很重要，但注意力不够，最终导致对环境的理解和影响达到新的水平。 与传染病作斗争。

ABSTRACT The sulfur-containing amino acid, L-cysteine, is indispensable for pathogen virulence and survival. Sulfur for biosyn- thesis is largely derived from the assimilation of inorganic sulfate by microorganisms. Genes required for sulfate as- similation are essential to pathogen survival but absent in the human genome and, therefore, represent potential new targets for therapeutic intervention. In the previous funding cycle, we defined key features in the catalytic cycle of sulfonucleotide reductases (SRs), enzymes that catalyze the first committed step in sulfate reduction for de novo synthesis of cysteine and other reduced-sulfur containing biomolecules. Our studies provided fundamental insights into how thioredoxins (Trxs)–central antioxidant enzymes that maintain protein thiols in their reduced state– recognize their cellular targets. We learned that the iron-sulfur cluster in APS reductase (APSR) plays an essential role active-site preorganization and substrate activation, expanding knowledge on the catalytic activities of Fe-S pro- teins and on the divergent evolution of PAPS reductase (PAPR), which lacks this cofactor. These insights led to molecules that target APSR in a new way, via interaction with the iron-sulfur metallocenter. We also discovered first- in-class inhibitors of sulfate reduction that exhibit potent bactericidal activity against drug-resistant clinical isolates of M. tuberculosis. Given the importance of SRs in pathogen oxidative stress resistance and virulence, we propose here to address research areas where the biggest open questions and greatest unmet needs remain: well-validated chemical probes that acutely inhibitor essential steps in microbial reductive sulfate assimilation (Aim 1); defining the non-redundant functions of pathogenic Trxs to facilitate sulfur reduction and cope with redox stress (Aim 2); and ad- vancing knowledge in trafficking and delivery of reactive sulfur species (RSS) for microbial cysteine biosynthesis. Collectively, the experiments in this renewal application will provide fundamental information on one of the most under-studied metabolic chemistries–reductive sulfate assimilation–in the context of one of the most devastat- ing pathogens–M. tuberculosis. We have established outstanding collaborations, with a proven track record of productivity, in order to support these efforts and ensure timely completion. Our efforts will address areas cen- tral to role of sulfur metabolism in pathogen oxidative stress resistance and virulence that have not received suf- ficient attention despite their importance, ultimately leading to a new level of understanding and leverage in the battle against infectious disease.