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）锚定的蛋白质（例如叶酸受体，prion蛋白和神经细胞粘附分子（NCAM））的生物合成至关重要，对正常细胞生长至关重要，并且对人类癌症以及属于遗传学的数量均受到扰动。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，以及阵发性夜间血红蛋白尿（PNH），这是一种获得的溶血性疾病，在多能血肿的人类干细胞中是由GPI生物合成的第一步引起的溶血性疾病。 GPI生物合成的遗传废除导致哺乳动物的胚胎致死性。 GPI组装途径已被证实是原生动物和真菌疾病的治疗靶点，包括非洲卧铺疾病和危及生命的机会性真菌感染，困扰着免疫受损的人。该提案的目的是了解GPI-蛋白的生物合成的各个方面，其总体目的是制定操纵和控制GPI途径的策略。这种努力是抗疾病和抗真菌药物的发展，以及癌症和遗传疾病的治疗，其中GPI锚定蛋白起关键的作用。我们对GPI蛋白质生物合成的两个至关重要的步骤感兴趣：GPI锚定在蛋白质上，以及跨内质网（ER）膜的GPI脂质的神秘触发器。 GPI跨成绩酶（GPIT）是连接GPI锚定在蛋白质上的酶，是一种鲜为人知的膜结合的杂型五体络合物。编码三个GPIT亚基的基因最近被鉴定为癌基因，从而提高了GPI途径可能为人类癌症治疗提供新的靶标的可能性。我们的目的是定义GPIT许多亚基的功能作用；我们对我们假设是该复合物的GPI结合元素的癌蛋白PIG-U和GAA1特别感兴趣。我们还建议对锥虫GPIT进行分析，从长远来看，该分析将产生与设计相关的产量结果，这些试剂可用于选择性地阻断锚定锥虫原生动物中的必需GPI，而这些寄生虫则是其哺乳动物宿主中的居民。我们的第二个目的是鉴定GPI Flippase，这是GPI组装过程中ER膜上转移（翻转）GPI脂质中间体所需的新型转运蛋白。 GPI翻转是GPI组装中的强制性步骤，也是唯一在生化和遗传上尚待定义的唯一一个步骤。由于未在功能上识别任何类型的ER脂质Flippase，因此我们识别GPI Flippase的目的不仅会特别有助于对GPI组装的理解，而且还将提供有关ER中脂质易位事件的基本信息。