STRUCTURE-FUNCTION RELATIONSHIPS OF PROTECTIN(CD59)

保护素(CD59)的结构与功能关系

• 批准号: 3366991
• 负责人: WENDELL F ROSSE
• 金额: $ 17.91万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-01 至 1996-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3366991
• 关键词: CD antigens; CHO cells; binding proteins; complement; complement inhibitors; complement pathway; complementary DNA; fusion gene; gene expression; genetic library; human tissue; lymphocyte; membrane proteins; molecular site; monoclonal antibody; nucleic acid sequence; paroxysmal nocturnal hemoglobinuria; phosphatidylinositols; polymerization; protein structure function; site directed mutagenesis; tissue /cell culture; transfection

CD59 (protectin, membrane inhibitor of reactive lysis [MIRL]) is a glycan- phosphatidylinositol (GPI) linked protein that is crucial in the control of complement activation by inhibiting the polymerization of C9. It is particularly important for cells of the blood and its lack in blood cells in paroxysmal nocturnal hemoglobinuria is the chief cause of hemolysis and perhaps thrombosis. Using a cDNA for CD59, we plan to characterize the gene structure of the molecule. We plan to determine structure-function relationships by replacing the GPI anchor with a transmembrane anchor and to vary the length of the cytoplasmic tail in order to determine the effect on its biological function. Using site-directed mutagenesis, we plan to determine which of the 11 cysteines in the molecule are important for function. We will replace segments of the molecule to determine the functional sites. We will examine the effect of altering the carboxyl end of the coded molecule on the fixation to the GPI anchor; specifically, we will mutagenize the three asparagine groups as well as specific residues within the hydrophobic region of the molecule. Finally, we will exchange this region with that of CD55 to determine if the hydrophobic region is protein specific. With these studies, we will have an increased understanding of the genetics and structure-function relationships of this important protein.

CD 59（protectin，反应性裂解的膜抑制剂[MIRL]）是一种聚糖- 磷脂酰肌醇（GPI）连接的蛋白质，这是至关重要的控制 通过抑制C9的聚合来激活补体。 是 对血液细胞特别重要， 阵发性睡眠性血红蛋白尿症是溶血的主要原因， 可能是血栓 使用CD 59的cDNA，我们计划表征 分子的基因结构。 我们计划确定结构-功能 通过用跨膜锚代替GPI锚来改变关系， 改变胞质尾的长度，以确定 它的生物学功能。 利用定点突变，我们计划 确定分子中的11个半胱氨酸中的哪一个对于 功能 我们将替换分子的片段以确定 功能性网站 我们将研究改变羧基末端的影响 在GPI锚的固定上的编码分子;具体地说，我们 将诱变三个天冬酰胺基团以及特定的残基 在分子的疏水区域内。 最后，我们将交换 将该区域与CD 55的区域进行比较，以确定疏水区域是否 蛋白质特异性。 通过这些研究，我们将增加 了解这种基因的遗传学和结构-功能关系， 重要蛋白质