ROLE OF WEAKLY BOUND CROSSBRIDGES IN MUSCLE CONTRACTION

弱束缚横桥在肌肉收缩中的作用

• 批准号: 3160942
• 负责人: JOSEPH M CHALOVICH
• 金额: $ 7.5万
• 依托单位: EAST CAROLINA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-04-01 至 1995-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3160942
• 关键词: X ray crystallography; actins; biochemistry; calcium; caldesmon; cell type; chemical kinetics; chickens; crosslink; drug design /synthesis /production; ionic strengths; laboratory rabbit; muscle contraction; myosins; single cell analysis; smooth muscle; stop flow technique; striated muscles; temperature; troponin; turkeys

The human muscle system contains three muscle types: cardiac muscle, skeletal muscle for physical stability and movement, and smooth muscle in the vasculature, digestive, and reproductive systems. To understand disorders affecting particular muscle types it is necessary to understand mechanisms both of contraction of these muscles and of regulation of contraction. Pharmacologic intervention depends on the ability to selectively modify the contractility of each muscle type. To this end we have been studying the regulation of contraction of both smooth and striated (cardiac and skeletal) muscles. We have shown earlier that the Ca2+ regulation of striated muscle, through the proteins troponin and tropomyosin, does not occur by a weakening of the binding of myosin-ATP to actin-tropomyosin-troponin. Rather, in the absence of Ca2+, the rate of a process, presumably product release, is inhibited. In contrast, it has been shown that the binding of myosin to actin-tropomyosin-troponin, in the absence of ATP, is Ca2+ dependent. These data are consistent with a model of contraction where both myosin makes the transition from a weakly bound state (containing bound ATP or ADP + Pi) to a strongly bound state (containing bound ADP). This transition cannot occur at a significant rate unless actin also makes a transition from the inactive state to the active state. We now propose to use a multidisciplinary approach to test this hypothesis and to further characterize the duality of states of actin and myosin. We have shown that the smooth muscle protein, caldesmon, does inhibit the binding of myosin-ATP to actin. This protein is an excellent probe of myosin crossbridge binding and can be used to study the function of the weakly bound state of myosin. We will use binding studies and stopped-flow kinetic studies to investigate the properties of weakly bound myosin crossbridges. These biochemical studies will be accompanied by mechanical and X-ray diffraction measurements of single muscle fibers to determine the importance of weakly bound crossbridges in contraction and in regulation. We will also use the muscle fiber model to study possible regulatory roles of caldesmon and a newly discovered protein, calponin. Finally, we will make a detailed study possible regulatory roles of caldesmon and a newly discovered protein, calponin. Finally, we will make a detailed study of the binding of myosin to actin in the presence of ATPgammaS to test a recent challenge to our hypothesis that regulation of striated muscle occurs at the transition from the weak binding states to the strong crossbridge states.

人类肌肉系统包含三种肌肉类型：心肌、 用于身体稳定和运动的骨骼肌，以及 血管系统、消化系统和生殖系统。要理解 影响特定肌肉类型的疾病有必要了解 这些肌肉的收缩和调节的机制 收缩。药物干预依赖于 有选择地改变每种肌肉类型的收缩能力。为此，我们 一直以来都在研究平滑和收缩的调节 横纹肌(心脏和骨骼)。我们早些时候已经表明， 钙离子通过肌钙蛋白和肌钙蛋白对横纹肌的调节 原肌球蛋白，不是由于肌球蛋白-ATP结合减弱而产生的 肌动蛋白原肌球蛋白肌钙蛋白。相反，在没有钙离子的情况下， 过程，大概是产品的释放，是被禁止的。相比之下，它有 已证明肌球蛋白与肌动蛋白-原肌球蛋白-肌钙蛋白的结合，在 缺乏ATP，是依赖于钙离子的。这些数据与一个模型一致 收缩，其中两个肌球蛋白都从弱结合过渡 状态(包含结合的ATP或ADP+PI)到强结合状态 (包含结合的ADP)。这种转变不可能以显著的速度发生。 除非肌动蛋白也从非活动状态转换为活动状态 州政府。我们现在建议使用多学科方法来测试这一点 并进一步刻画肌动蛋白和肌动蛋白的二元性。 肌球蛋白。我们已经证明，平滑肌蛋白钙调蛋白具有 抑制肌球蛋白-ATP与肌动蛋白的结合。这种蛋白质是一种极好的 肌球蛋白交叉桥结合探针可用于研究其功能 肌球蛋白的弱结合状态。我们将使用约束性研究和 停流动力学研究弱结合态的性质 肌球蛋白交叉连接。这些生化研究将伴随着 单根肌纤维的力学和X射线衍射测量 确定弱结合交叉桥在收缩和收缩中的重要性 监管。我们还将使用肌肉纤维模型来研究可能 钙调蛋白和一种新发现的蛋白质Calponin的调节作用。 最后，我们将对可能的监管角色进行详细研究 钙蛋白和一种新发现的蛋白质，钙蛋白。最后，我们将使 肌球蛋白与肌动蛋白结合的详细研究 ATPGammaS将测试最近对我们的假设的挑战，即对 横纹肌发生在从弱结合状态到 强大的交叉桥州。