MECHANISM OF CALCIUM OVERLOADING

钙超载的机制

• 批准号: 3356326
• 负责人: TUAN H KUO
• 金额: $ 12.29万
• 依托单位: WAYNE STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-07-01 至 1991-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3356326
• 关键词: adenosinetriphosphatase; affinity chromatography; calcium; calcium binding protein; calcium channel; calcium channel blockers; calmodulin; coronary disorder; disease /disorder model; enzyme structure; gas chromatography; hamsters; heart function; heart pharmacology; ionophores; liposomes; membrane lipids; membrane permeability; necrosis; phospholipids; sarcolemma; thin layer chromatography

The long-term goal of this project is to study the role of calcium transmembrane flux in maintaining normal cardiac structure and function. The spontaneous myocardial necrosis in hereditary cardiomyopathic Syrian hamster is accompanied by intracellular myocardial calcium overload. Based on preliminary findings suggesting depressed activity in the cardiac sarcolemmal Ca2+ pumping ATPase together with evidence of alteration in sarcolemmal membrane lipids in the myopathic hamster, we hypothesize that defects in this enzyme or in its regulation lead to decreased rate of calcium efflux at the plasma membrane level, resulting in calcium overload and then myocytolysis. In order to test this hypothesis, we propose: 1. To study alterations in both Ca2+ fluxes and plasma membrane lipids during the development of cardiomyopathy. Ca2+ efflux will be assessed by measuring the sarcolemmal Ca2+ pumping ATPase activity. Ca2+ influx through the voltage dependent Ca2+ channel will be assessed by using ligand binding assay. Membrane phospholipid and fatty acid composition will be analyzed by thin layer chromatography and gas-liquid chromatography. 2. To test for structural and functional defects in Ca2+ pumping ATPase. The enzyme will be purified by Calmodulin affinity chromatography and its biochemical characteristics determined. In addition, the kinetic properties of the enzyme as well as the effects of phosphatidylinositol lipids or calmodulin will be studied. The alterations in protein sequence will be studied by peptide mapping analysis using V8 protease and Cleveland digestion method. 3. To test for abnormal regulation of the Ca2+ pumping ATPase by membrane lipids. Ca2+ transport in reconstituted liposomes will be studied by using purified ATPase from human erythrocytes and sarcolemmal membrane lipids from both control and myopathic hamsters. Thus this project is not only relevant to the study of pathogenesis of cardiomyopathy but also may contribute to the understanding of the molecular properties of the plasma membrane calcium pump.

该项目的长期目标是研究钙的作用