MOLECULAR AND RED CELL DETERMINANTS OF SICKLE CELL DISEASE

镰状细胞病的分子和红细胞决定因素

• 批准号: 6606073
• 负责人: ROBERT M BOOKCHIN
• 金额: $ 22.47万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6606073
• 关键词: cell morphology; cell water; cellular pathology; clinical research; crosslink; erythrocyte membrane; erythrocytes; hemoglobin Ss; human subject; molecular pathology; oxygen transport; protein localization; protein structure function; recombinant proteins; sickle cell anemia; synthetic protein

DESCRIPTION (provided by applicant): This project brings the work and expertise of this laboratory on the molecular and cellular pathophysiology of sickle cell disease into collaboration with four other laboratories, each expert in different areas, with the common goal of a detailed description of the structure, interactions and kinetics of assembly of the deoxy-HB S polymers, fibers and gel and the behavior of Hb S in the red cell. We emphasize these objectives: 1) To characterize the interaction sites of the deoxy-Hb S polymers, fibers and gels, with particular attention to important sites susceptible to specific modification, by semisynthetic approaches and direct chemical modification; to the specific cis or trans role of specific residues at different structural levels of the polymer; and how non-S Hbs participate in polymerization; 2) to correlate our micro excluded volume assay of the polymer solubility, the dextran-Csat with Ferrone?s new micro-kinetic assay of polymerization, to permit thermodynamic analyses with this method; 3) to examine further the role of the polymer water compartment (PWC) on the redistribution of 2,3- DPG, ATP, RBC enzymes and small molecules which may affect both the polymerization process and its effects on cell metabolism; 4) to test for conformational changes in deoxy-Hb S polymerized in different conditions, to explain the observed exclusion of DPG, and solubility-lowering effect if IHP 4)to investigate the mechanisms of intracellular instability of Hb S, which generates hemichromes and oxidative membrane damage 5) to assess the role of normal or abnormal (SS-modified) components of the RBC membrane on the kinetics and assembly of intracellular polymer. Each recombinant, modified, and/or cross-linked hybrid tetramer Hb designed specifically to test the roles of individual sites in different levels of polymer and gel structure (Manning, Acharya), will be examined by our micro-assay of equilibrium solubility (dextran-Csat) and correlated with the micro-kinetic assay (Ferrone). Selected species will be examined by Joseph's electron microscopic methods, and by Briehl's methods of direct visualization of fiber growth and assembly. We share the goal of a detailed characterization of the structure and assembly mechanisms of the deoxy-Hb S polymer and gel, in solution and red cells, with the clinical aim of facilitating the design of specific therapeutic agents for sickle cell disease, and the basic aim of advancing a full model description of the mechanism and osmotic effects of protein polymerization and depolymerization in biological systems.

描述（由申请人提供）： 该项目带来了该实验室在分子和细胞上的工作和专业知识 镰状细胞疾病的病理生理学与其他四个实验室合作，每个实验室 在不同领域的专家，其共同目标是对结构的详细描述， 脱氧-Hb S聚合物，纤维和凝胶组装的相互作用和动力学 红细胞中HB S的行为。我们强调这些目标：1）表征 脱氧-Hb的聚合物，纤维和凝胶的相互作用位点，特别关注 重要的站点通过半合成方法和直接进行特定修饰的特定修饰。 化学修饰；特定的顺式或特定残基在不同结构上的trans作用 聚合物的水平；以及非S HBS如何参与聚合； 2）与我们的微观相关 排除了聚合物溶解度的体积测定，即具有Ferrone的新型微型运动测定的聚合物的右旋溶解度，以允许使用这种方法进行热力学分析； 3）到 进一步研究聚合物水室（PWC）对2,3-重新分布的作用 DPG，ATP，RBC酶和小分子可能影响两种聚合 过程及其对细胞代谢的影响； 4）测试脱氧-Hb s的构象变化 在不同条件下聚合，以解释IHP 4）研究HB S细胞内不稳定性的机制，从而解释观察到的DPG的排除，并降低溶解度的效果，从而产生半色膜和氧化膜损伤5），以评估正常或异常（SS-Modified）组成的作用（ss-Modified）组成的作用（ss-modified）的作用（ss-modified）embc embc embc embc embc embrane embrane embrane embrane embrane embrane embrane embrane embrane embrane son的作用 细胞内聚合物。每个重组，修饰和/或交联的混合四聚体HB 专门设计用于测试不同级别的聚合物和凝胶中各个位点的作用 结构（Manning，Acharya）将通过我们的平衡溶解度进行检查 （葡萄糖-CSAT），并与微型运动测定法（Ferrone）相关。选定的物种将是 由约瑟夫的电子显微镜方法和布里尔的直接方法检查 纤维生长和组装的可视化。我们分享了详细表征的目标 溶液和红色的脱氧-Hb S聚合物和凝胶的结构和组装机制 细胞，临床目的促进镰状细胞的特定治疗剂的设计 疾病，以及推进机制的完整模型描述的基本目的 蛋白质聚合和解聚在生物系统中的渗透作用。