NMR STRUCTURES OF PLATELET FACTOR 4 & B-THROMBOGLOBULIN

血小板因子 4 的 NMR 结构

• 批准号: 3361742
• 负责人: KEVIN H. MAYO
• 金额: $ 10.42万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-04-01 至 1993-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3361742
• 关键词: acidity /alkalinity; beta globulin; binding proteins; chemical binding; computer simulation; conformation; heparin; high performance liquid chromatography; human tissue; monomer; nuclear magnetic resonance spectroscopy; physical model; platelet factor 4; protein structure; protein structure function; solutions

Platelet factor (PF4) and B-thromboglobulin-related proteins (BTG) are low- molecular weight heparin binding proteins generally considered to be platelet specific and are involved in blood clotting, wound healing, tissue repair and cell proliferation. PF4 is known to strongly bind heparin thereby inhibiting the formation of thrombin-antithrombin III complexes at vascular sites. Other physiological activities of PF4 include stimulation of fibroblast attachment to the substrate, stimulation of histamine release from human basophils, chemotactic activity, and potentiation of platelet aggregation. While BTG is about 50% sequentially homologous to PF4, its anti-heparin activity is less, and it possesses significant mitogenic activity in certain cell types and inhibits the catalytic effect heparin on Factor Xa neutralization by anti-thrombin IIIa more than does PF4. Although the structures of PF4 and BTG are considered critical to biological function, little is known about their structure-function, relationships, about how their conformations are related, or about any specific structural domain(s) essential for interaction with heparin or glycosaminoglycans in general. The immediate goal of this project is to elucidate the solution structures of PF4 and low affinity-Pf4 (LA-PF4, the parent protein of the more commonly known BTG), and the long range goal is to identity the protein- heparin binding domain(s). Comparison of PF4 and BTG structures will give initial insight into this latter point. The primary technique to be used towards these goals is high-resolution, two-dimensional proton NMR spectroscopy in combination with computer modeling studies. By using COSY- like and NOESY NMR experiments, sequence-specific proton resonance assignments will be made for PF4 and BTG at low ph under conditions where these proteins exist as monomers (7,800 daltons). NOESY experiments will then be used to establish distance tetrameric structures at low ph than at more physiological conditions will be investigated by NMR methods just described. The tight association of four, tetrahedral-related, identical monomers makes structural elucidation of the 32 kD tetramer feasible by NMR methods.

血小板因子(PF4)和B-血栓球蛋白相关蛋白(BTG)低- 分子量肝素结合蛋白通常被认为是 血小板特异性，参与凝血、伤口愈合、组织 修复和细胞增殖。已知PF4与肝素强结合 从而抑制凝血酶-抗凝血酶III复合体的形成 血管部位。PF4的其他生理活性包括刺激 成纤维细胞附着在基质上，刺激组胺释放 来自人类嗜碱性粒细胞、趋化活性和增强血小板 聚合。虽然BTG与PF4的序列同源性约为50%，但其 抗肝素活性较低，具有显著的促有丝分裂作用。 在某些细胞类型中的活性，并抑制肝素对 抗凝血酶IIIa比PF4更能中和凝血因子Xa。虽然 PF4和BTG的结构被认为是生物学上的关键 功能，人们对它们的结构-功能，关系， 关于它们的构象是如何相关的，或者关于任何特定的结构 与肝素或糖胺聚糖相互作用所必需的结构域(S) 将军。 这个项目的直接目标是阐明溶液的结构 的PF4和低亲和力的-PF4(LA-PF4，母体蛋白更多 通常被称为BTG)，长期目标是识别蛋白质- 肝素结合区(S)。将对PF4和BTG结构进行比较 对后一点的初步洞察。要使用的主要技术 朝向这些目标的是高分辨率的二维质子核磁共振 光谱学与计算机模拟研究相结合。通过使用COSY- Like和NOESY核磁共振实验，序列特定的质子共振 在以下条件下，将在低ph值条件下对PF4和BTG进行分配 这些蛋白质以单体形式存在(7800道尔顿)。NOESY实验将 然后用来在低于pH的条件下建立距离四聚体结构 更多的生理状况将通过核磁共振方法进行研究 描述。四个与四面体相关的、完全相同的紧密联系 单体使32kD四聚体的核磁共振结构解析成为可能 方法：研究方法。