MOLECULAR BIOLOGY OF UNIQUE WALL LIPIDS OF MYCOBACTERIA

描述（改编自申请人摘要）： 与艾滋病有关的结核病和其他分枝杆菌感染 威胁着全世界的人类健康。 耐药结核病的出现 发现新的药物靶点的关键。 分枝杆菌细胞壁含有一种 含有多种类型的多甲基支链脂肪酸的各种脂质 是致病分枝杆菌所特有的 这些脂质被认为 在病原体逃避防御的能力中发挥重要作用 主持人的反应。 这些独特的脂质的生物合成， 多甲基分支细胞壁脂质可以提供新的理想目标， 抗分枝杆菌药物。 为了阐明参与的机制， 这些脂质的生物合成，并测试其可能的功能， 宿主-病原体相互作用，我们提出以下具体目标：1） 确定ORF 2表达是否是从 真菌蜡酸合成酶转化为苯硫代蜡醇和苯硫代酚的研究 orJ 2基因的破坏和对 真菌蜡基脂质生物合成。 如果ORF 2被发现是必要的，我们将 阐明其参与真菌蜡糖基脂质合成的机制 并确定ORF 2的结构。 2)阐明短链的性质 通过对该基因进行克隆和测序， 活动站点域组织和子单元组成。 3)击倒对手 在M.结核 并确定其生化和功能后果。 4)纯化亚 新发现的支链脂肪酸合酶，鉴定产生的产物， 在基因组数据中寻找编码合酶的基因的身份 库使用蛋白质序列，并确定生化和 基因敲除的功能性后果。 5)确定 目的研究M.结核 其产物的表征以M. smegalloy，and by 确定其生物化学和功能后果， 破坏 6)确定Class 2（msl 3、msl 4和msl 5）的功能 和其他mas样基因。结核病基因组，通过表征 其产物以M.，并通过测定 生物化学和功能的后果。 7)确定 是否缺乏特定的脂质引起的上述基因破坏影响 宿主-病原体相互作用和毒力。 结果将有助于识别 细胞壁脂质对发病机制至关重要，可能是合适的靶点 新的抗分枝杆菌药物。

DESCRIPTION (Adapted from the applicant's abstract): Resurgence of tuberculosis and other mycobacterial infections associated with AIDS threaten human health world-wide. Emergence of drug resistant TB makes it critical to discover new drug targets. Mycobacterial cell walls contain a variety of lipids with many types of multiple methyl-branched fatty acids that are unique to pathogenic mycobacteria. These lipids are thought to play important roles in the ability of the pathogens to evade the defense reactions of the host. Biosynthesis of these unique lipids containing multiple-methyl branched cell wall lipids could offer ideal targets for new antimycobacterial drugs. To elucidate the mechanisms involved in the biosynthesis of these lipids and to test for their possible functions in host-pathogen interaction, we propose the following specific aims: 1) Determine whether ORF2 expression is required for mycocerosyl transfer from mycocerosic acid synthase to phthiocerol and phenolphthiocerol in vivo by disruption of the orJ2 gene and examination of the consequence on mycocerosyl lipid biosynthesis. If ORF2 is found to be necessary, we will elucidate the mechanism of its participation in mycocerosyl lipid synthesis and determine the structure of ORF2. 2) Elucidate the nature of short-chain mycocerosic acid synthase by cloning and sequencing the gene to identify the active site domain organization and subunit composition. 3) Knock-out the gene that encodes short-chain mycocerosic acid synthase in M. tuberculosis and determine the biochemical and functional consequences. 4) Purify a newly-found branched fatty acid synthase, identify the products generated, seek the identity of the gene that encodes the synthase in the genomic data bank using the protein sequence, and determine the biochemical and functional consequences of knocking-out this gene. 5) Determine the functions of Class 1 mas-like genes, msl1 and msl2, of M. tuberculosis by characterization of their products expressed in M. smegmatis, and by determination of the biochemical and functional consequences of their disruption. 6) Determine the functions of Class 2 (msl3, msl4, and msl5) and other mas-like genes in M. tuberculosis genome, by characterization of their products expressed in M. smegmatis, and by determination of the biochemical and functional consequences of their disruption. 7) Determine whether lack of specific lipids caused by the above gene-disruptions affects host-pathogen interaction and virulence. The results will help identify cell wall lipids critical for pathogenesis that might be suitable targets for new antimycobacterial drugs.