GLYCOSYLATION MUTANTS OF LEISHMANIA

利什曼原虫糖基化突变体

• 批准号: 2672053
• 负责人: Salvatore J Turco
• 金额: $ 49.01万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-07-01 至 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2672053
• 关键词: Leishmania donovani; disaccharides; enzyme activity; flow cytometry; gene expression; gene mutation; gene targeting; genetic regulation; glycolipids; glycoprotein biosynthesis; glycosylation; hamsters; host organism interaction; intracellular parasitism; laboratory mouse; leishmaniasis; membrane proteins; molecular cloning; northern blottings; nucleic acid sequence; phosphatidylinositols; polymerase chain reaction; protein structure function; southern blotting; transposon /insertion element; virulence

A striking feature of Leishmania parasites is their ability to survive in hostile environments encountered throughout their life cycle. Lipophosphoglycan (LPG) is the major cell surface glycoconjugate of all Leishmania promastigotes and plays a key role in the survival of the parasites. Prior studies have characterized the LPG from L. donovani as a polymer of repeating disaccharide units of [-6Gal(beta1,4)Man 1-PO4], attached via a glycan core to a novel lyso-alkylphosphatidyl inositol anchor. One powerful approach for establishing the biosynthesis and function of LPG is the identification of genes encoding proteins required for its synthesis, and the development of mutants defective in one or more aspects of LPG biosynthesis. We have developed effective methods for creating lpg- mutants, and for the identification of LPG biosynthetic genes capable of rescuing the defect in these mutants. With these, 5 different genes involved in LPG biosynthesis have been identified, and their biochemical and biological roles established. Our studies have provided new insight into general mechanisms of glycoconjugate synthesis relevant to many eukaryotes, and several of the encoded proteins show promise as targets for future chemotherapeutic efforts. The ultimate goal is to develop an comprehensive understanding of the genes involved in LPG biosynthesis, the organization of this complex and important biochemical pathway including its cellular compartmentalization, and the biological function of LPG. Our specific aims are: l) To identify new lpg- mutants and identify the defective genes in Leishmania donovani. We will also explore an exciting new approach, using transposable elements within the parasite, to tag and inactivate LPG genes. 2) To determine the biochemical consequences of each lpg- mutation on the biosynthesis of LPG, and molecules bearing LPG-related elements, such as the disaccharide phosphate repeating unit and GPI anchors. 3) To determine the biochemical role of the protein(s) encoded by each LPG gene. 4) To utilize carefully chosen lpg- mutants and controls to establish the role of each gene in the parasite infectious cycle.

利什曼原虫的一个显著特征是它们的生存能力 在整个生命周期中遇到的恶劣环境中。 脂磷酸聚糖（LPG）是所有细胞的主要细胞表面糖缀合物。 利什曼原虫前鞭毛体，并在生存的关键作用， 寄生虫先前的研究已经表征了来自L.多诺瓦尼阿斯 [-6Gal（beta1，4）Man 1-PO 4]的重复二糖单元的聚合物， 通过聚糖核心与新型溶血烷基磷脂酰肌醇连接 锚。一种建立生物合成的有效方法， LPG的功能是识别编码蛋白质的基因 所需的合成，和突变体的发展缺陷， LPG生物合成的一个或多个方面。 我们已经开发出有效的方法来创造液化石油气突变体， LPG生物合成基因的鉴定能够拯救 这些突变体的缺陷。有了这些，5个不同的基因参与LPG 生物合成已被确定，其生物化学和生物 角色设定。我们的研究提供了新的见解， 与许多真核生物相关的糖缀合物合成机制，以及 一些编码的蛋白质显示出作为未来靶点的希望。 化疗的努力 最终目标是全面了解 基因参与LPG的生物合成，这种复合体的组织， 重要的生化途径，包括其细胞 区室化和LPG的生物学功能。我们的具体 目标是： l）鉴定新的lpg-突变体并鉴定lpg-突变体中的缺陷基因。 杜氏利什曼原虫我们还将探索一种令人兴奋的新方法， 利用寄生虫体内的转座因子， LPG基因。 2)为了确定每一个lpg突变的生化后果， LPG的生物合成，以及携带LPG相关元素的分子， 例如磷酸双糖重复单元和GPI锚。 3)为了确定由每个编码的蛋白质的生物化学作用， LPG基因 4)利用精心挑选的液化石油气突变体和对照， 每个基因在寄生虫感染周期中的作用。