Biogenesis of lipoarabinomannan in mycobacteria

分枝杆菌中阿拉伯脂甘露聚糖的生物发生

• 批准号: 7054643
• 负责人: Patrick Joseph Brennan
• 金额: $ 35.69万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-05-01 至 2010-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7054643
• 关键词: Mycobacterium bovis; Mycobacterium smegmatis; Mycobacterium tuberculosis; bacterial genetics; carbohydrate biosynthesis; cell free system; genetic library; glycosyltransferase; lipopolysaccharides; monoclonal antibody; mutant; protein structure; transposon /insertion element

DESCRIPTION (provided by applicant): Phosphatidylinositol mannosides (PIM) and their multiglycosylated counterparts, lipomannan (LM) and lipoarabinomannan (LAM) are complex glycolipids/ lipoglycans that are found in the envelopes of all mycobacterial species. They are important immunomodulatory molecules in the course of tuberculosis and leprosy as well as key ligands promoting the entry of mycobacteria into phagocytic and non-phagocytic cells. Although much progress has been made over the last 15 years in deciphering the structures and structure-function relationships of these molecules, little is known about their biogenesis. The elucidation of their biosynthetic pathways, in addition to providing fundamental knowledge about the biochemistry of Mycobacterium tuberculosis, would lead to the discovery of essential enzymes that could represent crucial targets for novel anti-tuberculosis drugs. The availability of defined M. tuberculosis and M. bovis BCG mutants deficient in some aspects of PIM/ LM and LAM synthesis would also enable a precise measurement of the contribution of these molecules to the immunopathogenesis of tuberculosis and suggest new vaccine strategies against this disease. We propose to exploit the recent knowledge of the structures of LAM-like molecules produced by actinomycetes together with the availability of a growing number of genome sequences of lipoglycan-producing actinomycetes to perform genome comparison studies, identify glycosyltransferases involved in the biogenesis of PIM, LM and LAM and construct lipoglycan-deficient mutants of mycobacteria. Using the CS- 35 anti-LAM monoclonal antibody and other antibodies that we have developed, we also propose to screen transposon mutant libraries of M. tuberculosis, M. bovis BCG, and M. smegmatis for mutants defective in some aspects of LAM synthesis. Mutants will be biochemically characterized using conventional methods and novel analytical tools, cell-free assays will be developed for the newly identified biosynthetic enzymes and the three-dimensional structure of these proteins will be determined. Finally, the interactions of PIM/ LM/ LAM mutants with host cells and their immunomodulatory properties will be investigated.

描述（由申请人提供）：磷脂酰肌醇甘露糖苷（PIM）及其多糖基化对应物，LipoMannan（LM）和脂肪氨基氨基氨基氨基类（LAM）是复杂的糖脂/脂肪糖，是在所有mycobacterial种类的包裹中都发现的。它们是结核病和麻风病过程中的重要免疫调节分子，以及促进分枝杆菌进入吞噬细胞和非吞噬细胞的关键配体。尽管在过去的15年中取得了很大的进步，但在解释了这些分子的结构和结构 - 功能关系，但对它们的生物发生知之甚少。除了提供有关结核分枝杆菌生物化学的基本知识之外，还要阐明其生物合成途径，还将导致发现必需酶，这些酶可能代表新型抗链球菌药物的重要靶标。在PIM/ LM和LAM合成的某些方面缺乏定义的结核分枝杆菌和牛乳杆菌BCG突变体的可用性也将对这些分子对结核病的免疫发生的贡献进行精确的测量，并提出针对这种疾病的新疫苗策略。 我们建议利用放线菌产生的LIM样分子的结构，以及越来越多的脂肪糖糖促进性放线菌的基因组序列的可用性，以进行基因组比较研究，以鉴定糖基转移酶涉及PIM和LMCOMCOM和LAM cotcan-diccogcis of Pim的生物形成。使用CS-35抗LAM单克隆抗体和其他我们开发的抗体，我们还建议在LAM合成的某些方面筛选结核分枝杆菌，BOVIS BCG和SMEGMATIS的转骨突变库和Smegmatis的突变体有缺陷的突变体。将使用常规方法和新型分析工具对突变体进行生化表征，将为新鉴定的生物合成酶开发无细胞的测定，并确定这些蛋白质的三维结构。最后，将研究PIM/ LM/ LAM突变体与宿主细胞及其免疫调节特性的相互作用。