STRUCTURE/FUNCTION STUDIES OF VITAMIN D BINDING PROTEIN

维生素 D 结合蛋白的结构/功能研究

• 批准号: 2701102
• 负责人: RAHUL RAY
• 金额: $ 16.35万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-01-01 至 2001-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2701102
• 关键词: 1,25 dihydroxycholecalciferol; 25 hydroxycholecalciferol; X ray crystallography; affinity labeling; chemical binding; fatty acid binding protein; hormone binding protein; hormone regulation /control mechanism; macrophage activating factor; molecular site; osteoclast activating factor; point mutation; protein sequence; protein structure function; vitamin D; vitamin analog

DESCRIPTION (Adapted from the Applicant's Abstract): Vitamin D-binding protein (DBP) binds vitamin D and its metabolites as well as G-actin with high affinity and serves to transport the former to target tissues, and to scavenge the latter so as to prevent its polymerization during cell-injury. DBP also binds fatty acids, and enhances complement activation on neutrophil chemotaxis by binding to C5a des Arg. Additionally, DBP is known to be converted in vivo to a potent macrophage and osteoclast activating factor (DBP-MAF). The objectives of this proposal are to study various structure-functional aspects of DBP and DBP-MAF using a multiple-methods approach including chemical modification of the vitamin D sterol and fatty acid-binding domains of DBP by affinity/photoaffinity labeling, manipulations of the DBP-gene to express mutants and segments of DBP and their functional characterization, determination of the 3-D structures of DBP and DBP-actin complex by X-ray crystallography, and macrophage-superoxide production and bone resorption assays (for DBP-MAF activities). Multi-tasking nature of DBP is directly related to specific recognition/binding of various ligands by DBP. Such processes are strongly influenced by the structure of the protein, particularly the 3-D structures of the domains of the protein that are responsible for such recognition and binding. Hence, these studies will provide information about the importance of independent domains in the multi-domained structure of DBP, possible sharing of a common binding pocket by various endogenous ligands, degree of cross-talk among various domains, interdependence among various ligands, and influence of ligand-binding on DBP-MAF activities. The role of DBP is crucial in the metabolic activation of 1,25(OH)2D3, the calciotropic hormone, and its tissue-specific delivery, so that the latter can be responsible in calcium and phosphorus homeostasis, regulation of growth and maturity of cells, antiproliferation of malignant cells, and immunoregulation. Interaction of DBP with G-actin is important in the prevention of actin-polymerization, and clogging of arteries during cellular injury. On the other hand, macrophage-activation by DBP-MAF has stressed the possible immunoregulatory property of DBP. Furthermore, osteoclast-activation (by DBP-MAF) has raised the possibility that this cytokine may be involved in inflammatory joint diseases such as osteoarthritis, rheumatoid arthritis, periodontal diseases, etc. The proposed structure-function studies should be valuable in evaluating multiple functions and their possible physiological implications of DBP and DBP-MAF.

描述（改编自申请人的摘要）：维生素D结合 蛋白质（DBP）结合维生素D及其代谢产物以及G-肌动蛋白与 高亲和力，并将前者运送到靶向组织，并将其运送到 清除后者，以防止其在细胞伤害期间的聚合。 DBP还结合脂肪酸，并增强对中性粒细胞的补体激活 通过与C5a des arg结合趋化性。此外，已知DBP是 在体内转换为有效的巨噬细胞和破骨细胞激活因子 （DBP-MAF）。 该提案的目标是研究各种结构功能 DBP和DBP-MAF的各个方面使用多个方法方法包括 维生素D固醇和脂肪酸结合域的化学修饰 通过亲和力/光性标签的DBP，对DBP-GENE的操作 表达DBP的突变体和段及其功能表征， 通过X射线测定DBP和DBP-肌动蛋白复合物的3-D结构 晶体学和巨噬细胞蛋白氧化物的产生和骨吸收 测定（用于DBP-MAF活动）。 DBP的多任务性质与特定 DBP对各种配体的识别/结合。 这样的过程很强烈 受蛋白质结构的影响，尤其是3-D结构 蛋白质领域负责这种识别和 结合。 因此，这些研究将提供有关重要性的信息 DBP多域结构中的独立域的可能 通过各种内源配体共享共同的结合口袋，程度 各个领域之间的串扰，各种配体之间的相互依赖性， 配体结合对DBP-MAF活动的影响。 DBP的作用在1,25（OH）2d3的代谢激活中至关重要 钙称激素激素及其组织特异性递送，以便后者 可以负责钙和磷稳态，调节 细胞的生长和成熟度，恶性细胞的抗增殖以及 免疫调节。 DBP与G-肌动蛋白的相互作用在 预防肌动蛋白聚合和细胞期间动脉堵塞 受伤。 另一方面，DBP-MAF的巨噬细胞激活已经强调 DBP可能的免疫调节特性。 此外， 破骨细胞激活（通过DBP-MAF）提高了这样的可能性 细胞因子可能参与炎症关节疾病，例如 骨关节炎，类风湿关节炎，牙周疾病等。 建议的结构功能研究对于评估应该很有价值 多个功能及其对DBP和 DBP-MAF。