Molecular Interactions of Fibrinolysis

纤溶的分子相互作用

• 批准号: 6871184
• 负责人: JOHN Y ANAGLI
• 金额: $ 32.18万
• 依托单位: HENRY FORD HEALTH SYSTEM
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 2008-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6871184
• 关键词: active sites; acylation; antibody neutralization test; chemical kinetics; conformation; enzyme mechanism; enzyme structure; enzyme substrate complex; fibrin; fibrinolysis; fluorescent dye /probe; fluorimetry; intermolecular interaction; molecular dynamics; monoclonal antibody; plasminogen; plasminogen activator; plasminogen activator inhibitors; protease inhibitor; protein structure function; serine proteinases; site directed mutagenesis; urokinase; vitronectin

DESCRIPTION (provided by applicant): The overall goal of this project is to elucidate, on a molecular level, the mechanisms of inhibitory control of fibrinolysis by plasminogen activator inhibitor-1 (PAI-1). The following three specific aims will be pursued: (1) Determine the structure/function, mechanisms of inhibition and target specificity for PAI-1 inhibition of serine proteases. This will involve identification of critical residues and specific protein regions which are key to PAI-1 function. (2) Neutralization of PAI-1 by monoclonal antibodies. We will investigate four types of antibodies which inactivate the PAI-1 inhibitory function. The first type accelerates the rate of spontaneous conversion of PAI-1 to a latent form: the second type competes with proteinases for binding to PAI-1; the third type retards the rate of reactive center loop (RCL) insertion: and the fourth type probably prevents the disordering of the proteinase active center by PAI-l after RCL insertion. (3) Identify structural determinants, mechanisms and functional aspects that regulate the molecular interactions of PAI-1 with vitronectin (Vn) and fibrin. Vitronectin is the key molecule involved in interaction of PAI-1 with cell surfaces and fibrin clots. We intend to study the cooperative interactions and conformational changes of PAI-1 and monomeric and polymeric Vn. Our experimental approach will involve site-directed mutagenesis, fluorescence techniques, and rapid reaction kinetics to observe the rates of formation and decay of transient intermediates. This project can provide a foundation at the molecular level for rational development of therapeutic agents to modulate PAI-1 activity.

描述（由申请人提供）：本项目的总体目标是 阐明，在分子水平上，抑制控制的机制， 纤溶酶原激活物抑制剂-1（派-1）的纤溶作用。以下三 具体目标如下：（1）确定结构/职能、机制 派-1抑制丝氨酸蛋白酶的抑制和靶特异性。 这将涉及关键残留物和特定蛋白质的鉴定 这些区域是派-1功能的关键。(2)派-1的中和 克隆抗体我们将研究四种类型的抗体， 派-1抑制功能。第一种是加快速度 派-1自发转化为潜伏形式：第二种类型竞争 与派-1结合的蛋白酶;第三种类型延缓了 反应性中心回路（RCL）插入：第四种类型可能会阻止 RCL插入后PAI-1对蛋白酶活性中心的紊乱。（三） 确定结构决定因素、机制和功能方面， 调节派-1与玻连蛋白（Vn）和纤维蛋白的分子相互作用。 玻连蛋白是派-1与细胞相互作用的关键分子 表面和纤维蛋白凝块。我们打算研究合作的相互作用， 派-1和单体和聚合物Vn的构象变化。 我们的实验方法将包括定点突变，荧光 技术和快速反应动力学，以观察形成速率， 瞬态中间体的衰变。该项目可以提供一个基础 合理开发治疗剂的分子水平 派-1活性。

项目成果

Saturation mutagenesis of the plasminogen activator inhibitor-1 reactive center.

纤溶酶原激活剂抑制剂-1 反应中心的饱和诱变。

• 发表时间: 1992
• 期刊: The Journal of biological chemistry
• 作者: Sherman,PM;Lawrence,DA;Yang,AY;Vandenberg,ET;Paielli,D;Olson,ST;Shore,JD;Ginsburg,D
• 通讯作者: Ginsburg,D

The acid stabilization of plasminogen activator inhibitor-1 depends on protonation of a single group that affects loop insertion into beta-sheet A.

纤溶酶原激活剂抑制剂-1 的酸稳定性取决于影响环插入 β-片层 A 的单个基团的质子化。

• DOI: 10.1074/jbc.270.46.27942
• 发表时间: 1995
• 期刊: The Journal of biological chemistry
• 作者: Kvassman,JO;Lawrence,DA;Shore,JD
• 通讯作者: Shore,JD

A fluorescent probe study of plasminogen activator inhibitor-1. Evidence for reactive center loop insertion and its role in the inhibitory mechanism.

纤溶酶原激活剂抑制剂1的荧光探针研究。

• DOI: 10.1074/jbc.270.10.5395
• 发表时间: 1995
• 期刊: The Journal of biological chemistry
• 作者: Shore,JD;Day,DE;Francis-Chmura,AM;Verhamme,I;Kvassman,J;Lawrence,DA;Ginsburg,D
• 通讯作者: Ginsburg,D

The use of fluorescent probes to characterize conformational changes in the interaction between vitronectin and plasminogen activator inhibitor-1.

使用荧光探针来表征玻连蛋白和纤溶酶原激活剂抑制剂-1 之间相互作用的构象变化。

• DOI: 10.1074/jbc.272.8.5112
• 发表时间: 1997
• 期刊: The Journal of biological chemistry
• 作者: Gibson,A;Baburaj,K;Day,DE;Verhamme,I;Shore,JD;Peterson,CB
• 通讯作者: Peterson,CB