Biosynthesis of Membrane Protein Glycolipid Anchors

膜蛋白糖脂锚的生物合成

• 批准号: 7938503
• 负责人: ANANT K MENON
• 金额: $ 19.8万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7938503
• 关键词: Abbreviations; Acetylcholinesterase; Acetylglucosamine; African Trypanosomiasis; Anabolism; Antifungal Agents; Benzophenones; Binding; Binding Proteins; Binding Sites; Biochemistry; Biological; Biological Assay; COOH-terminal signal transamidase; Carrier Proteins; Catalytic Domain; Chemicals; Complex; Conserved Sequence; Data; Defect; Detergents; Development; Disease; Dolichol; Elements; Embryo; Endoplasmic Reticulum; Enzymes; Ethanolamines; Event; Evolution; Face; Family suidae; GPI Membrane Anchors; Genes; Genetic; Glucosamine; Glycolipids; Glycosylphosphatidylinositol-anchor Biosynthesis Pathway; Glycosylphosphatidylinositols; Hemagglutinin; Hematopoietic; Hereditary Disease; Human; Individual; Inherited; Label; Life; Lipids; Liver; Malignant Neoplasms; Mammals; Mannose; Maps; Membrane; Membrane Proteins; Molecular; Mutagenesis; Mycoses; Neural Cell Adhesion Molecules; Normal Cell; Oncogene Proteins; Oncogenes; PI-Glycan; Parasites; Pathway interactions; Pharmaceutical Preparations; Phosphatidylinositols; Phospholipase C; Phospholipids; Play; Positioning Attribute; Prions; Procedures; Protein Biosynthesis; Proteins; Protozoa; Radiolabeled; Rattus; Reaction; Reagent; Recruitment Activity; Reporter; Research Personnel; Role; Seizures; Side; Structure; System; Testing; Triton X100; Trypanosoma; Trypanosoma brucei brucei; Venous Thrombosis; Work; base; cancer therapy; catalyst; cell growth; design; dolichyl-diphosphooligosaccharide - protein glycotransferase; fatty acid elongases; folate-binding protein; fungus; glycoprotein phospholipase D; human stem cells; innovation; interest; man; mutant; novel; phosphoethanolamine; programs; proteoliposomes; radiotracer; rat Piga protein; reconstitution; therapeutic target; transamidases

DESCRIPTION (provided by applicant): The biosynthesis of glycosylphosphatidylinositol (GPI)-anchored proteins (such as the folate receptor, prion protein, and the neural cell adhesion molecule (NCAM)) is critical for normal cell growth, and perturbed in human cancers as well as a number of genetic diseases. The latter include an inherited GPI deficiency characterized by venous thrombosis and seizures, and paroxysmal nocturnal hemoglobinuria (PNH), an acquired hemolytic disease that is caused by a defect in the first step of GPI biosynthesis in multipotent hematopoietic human stem cells. Genetic abrogation of GPI biosynthesis results in embryonic lethality in mammals. The GPI assembly pathway has been validated as a therapeutic target for protozoal and fungal diseases, including African sleeping sickness and the life-threatening opportunistic fungal infections that afflict immuno-compromised individuals. The aims of this proposal are to understand aspects of the biosynthesis of GPI-proteins with the overall objective of developing strategies to manipulate and control the GPI pathway. Such efforts are central to the development of anti-protozoal and anti-fungal drugs, as well as to the treatment of cancer and inherited diseases in which GPI-anchored proteins play a key part. We are interested in two critically important steps of GPI-protein biosynthesis: the attachment of a GPI anchor to protein and the enigmatic flip-flop of GPI lipids across the endoplasmic reticulum (ER) membrane. GPI transamidase (GPIT), the enzyme that attaches GPI anchors to protein, is a poorly understood membrane-bound hetero-pentameric complex. Genes encoding three of the GPIT subunits have been recently identified as oncogenes, raising the possibility that the GPI pathway may provide a novel target for human cancer treatment. Our aim is to define the functional role of GPIT's many subunits; we are especially interested in the oncoproteins PIG-U and GAA1 that we hypothesize to be the GPI binding elements of the complex. We also propose analyses of trypanosome GPIT that will, in the long term, yield results pertinent to the design of reagents that could be used to selectively block essential GPI anchoring in trypanosomatid protozoa while these parasites are resident in their mammalian hosts. Our second aim is to identify GPI flippase, the novel transporter that is required to translocate (flip) GPI lipid intermediates across the ER membrane during GPI assembly. GPI flipping is an obligatory step in GPI assembly and the only one that currently remains to be biochemically and genetically defined. Since no ER lipid flippase of any type has been functionally identified, our aim to identify the GPI flippase will not only contribute specifically to an understanding of GPI assembly but will also provide fundamental information on lipid translocation events in the ER in general.

描述（由申请人提供）：糖基磷脂酰肌醇（GPI）锚定蛋白（如叶酸受体、朊病毒蛋白和神经细胞粘附分子（NCAM））的生物合成对于正常细胞生长至关重要，并且在人类癌症以及许多遗传疾病中受到干扰。后者包括以静脉血栓形成和癫痫发作为特征的遗传性GPI缺乏症，以及阵发性睡眠性血红蛋白尿症（PNH），这是一种获得性溶血性疾病，由多能造血干细胞中GPI生物合成的第一步缺陷引起。GPI生物合成的遗传废除导致哺乳动物的胚胎致死。GPI组装途径已被验证为原生动物和真菌疾病的治疗靶点，包括非洲昏睡病和折磨免疫受损个体的危及生命的机会性真菌感染。本提案的目的是了解GPI蛋白生物合成的各个方面，总体目标是制定策略来操纵和控制GPI途径。这些努力对于抗原生动物和抗真菌药物的开发以及GPI锚定蛋白发挥关键作用的癌症和遗传性疾病的治疗至关重要。我们对GPI蛋白生物合成的两个关键步骤感兴趣：GPI锚与蛋白质的连接和GPI脂质穿过内质网（ER）膜的神秘触发器。GPI转酰胺酶（GPIT）是一种膜结合的异源五聚体复合物，它是将GPI锚定物连接到蛋白质上的酶。编码三个GPIT亚基的基因最近被鉴定为癌基因，这提高了GPI途径可能为人类癌症治疗提供新靶点的可能性。我们的目标是确定GPIT的许多亚基的功能作用，我们特别感兴趣的是癌蛋白PIG-U和GAA 1，我们假设是GPI的结合元件的复杂。我们还提出了锥虫GPIT的分析，将在长期内，产生相关的试剂，可用于选择性地阻止必要的GPI锚定在锥虫原虫，而这些寄生虫是居民在其哺乳动物宿主的设计结果。我们的第二个目标是确定GPI翻转酶，这是一种新型的转运蛋白，在GPI组装过程中，它需要将GPI脂质中间体转运（翻转）穿过ER膜。GPI翻转是GPI组装的一个必要步骤，也是目前唯一一个有待生物化学和遗传学定义的步骤。由于没有任何类型的ER脂质翻转酶的功能已被确定，我们的目标，以确定GPI翻转酶不仅有助于具体的GPI组装的理解，但也将提供基本信息的脂质易位事件在ER中一般。