Investigation of Mycobacterial GMC Oxidoreductase Rv3409c

描述（由申请人提供）：世界卫生组织估计，每年有200万人死于结核病，世界人口的30%受到感染。多重耐药微生物广泛存在，广泛耐药微生物正在出现。需要新的方法来对抗这些毒性和耐药生物体（MDR/XDR TB）。 Rv 3409 c的功能未知。它与细菌胆固醇氧化酶有远亲关系，在基因组测序期间被注释为胆固醇氧化酶。用纯化的蛋白质和转座子突变体进行的实验表明，Rv 3409 c不是胆固醇氧化酶。突变体具有菌落形态表型，脂质谱分析已鉴定突变体中存在的脂质不存在于野生型分枝杆菌中。 所提出的实验旨在表征Rv 3409 c。假设Rv 3409 c是糖脂氧化酶，并且该活性代表抗MDR-TB药物开发的新靶标。这项工作的目的是1）阐明Rv 3409 c突变体中独特或缺失的脂质结构和2）确定Rv 3409 c酶的动力学活性和底物特异性。 公共卫生相关性：拟议的研究符合NIAID的目标研究需求，该研究旨在开发针对MDR/XDR TB的新化疗药物。这些实验将为发现这种不存在于人类中的酶的功能提供基础，并指导新抗生素的开发，这些抗生素将有效对抗所有形式的结核病，包括MDR/XDR结核病。

DESCRIPTION (provided by applicant): The World Health Organization estimates that 2 million people die every year from tuberculosis and that 30% of the world's population is infected. Multi-drug resistant organisms are widespread and extensively drug resistant organisms are emerging. New approaches are required to combat these virulent and drug resistant organisms (MDR/XDR TB). The function of Rv3409c is unknown. It is distantly related to bacterial cholesterol oxidases and was annotated as a cholesterol oxidase during genome sequencing. Experiments with purified protein and transposon mutants revealed that Rv3409c is not a cholesterol oxidase. The mutant has a colony morphology phenotype and lipid profiling has identified lipids present in the mutant that are not present in wild-type mycobacteria. The proposed experiments are aimed at characterizing Rv3409c. The hypothesis is that Rv3409c is a glycolipid oxidase and that this activity represents a new target for anti-MDR-TB pharmaceutical development. The aim of this work is 1) to elucidate the structure of lipids unique or missing in the Rv3409c mutant and 2) to determine the kinetic activity and substrate specificity of the Rv3409c enzyme. PUBLIC HEALTH RELEVANCE: The proposed research fits within the targeted research needs of NIAID that seeks the development of new chemotherapeutic agents against MDR/XDR TB. These experiments will provide a basis for discovering the function of this enzyme that is not present in humans and guide development of new antibiotics that would be effective against all forms of TB including MDR/XDR TB.