TOPOGRAPHICAL STUDIES OF SMOOTH MUSCLE MYOSIN

将使用荧光技术来探测Smooth的拓扑结构 肌肉肌球蛋白。这些实验的主要目的是回答问题 关于调节平滑肌活动的分子机制。 荧光和/或发色化合物将特别用于 标记肌球蛋白活性部位，重链中的不同位置， 两种不同类型的轻链，以及杆子上的特定位置 分子的一部分。一种新的探头贴合方法 特别是指被激酶磷酸化的丝氨酸或苏氨酸 提出了一种新的解决方案。这种标记方法具有普遍应用的潜力 研究其他体系中磷酸化中心的结构方面 特别是被蛋白激酶磷酸化。荧光能量转移 技术将被用作光谱标尺来探测构象 可能对平滑肌作用机制很重要的转变 肌球蛋白轻链磷酸化调节。的影响 肌球蛋白光对调节轻链的磷酸化作用 这些距离上的链激酶或蛋白激酶C将被检测。 使用这些方法，假设头部和尾部的连接点 肌球蛋白分子在平滑调节机制中起关键作用 肌肉肌球蛋白将进行测试。这些信息将有助于 确定依赖于磷酸化的构象如何变化 体外观察与生理上重要的结构变化有关 在体内的肌球蛋白细丝内。

Fluorescence techniques will be used to probe the topology of smooth muscle myosin. The major goal of these experiments is to answer questions about the molecular mechanism of regulation of smooth muscle activity. Fluorescent and/or chromophoric compounds will be used to specifically label the myosin active site, various positions within the heavy chain, the two different kinds of light chains, and a specific site on the rod portion of the molecule. A novel new method of attaching probes specifically to serines or threonines that are phosphorylated by kinases is presented. This labeling method has potential to be generally useful to study the structural aspects of phosphorylated sites in other systems specifically phosphorylated by kinases. Fluorescence energy transfer techniques will be used as a spectroscopic ruler to probe conformational transitions which may be important to the mechanism of smooth muscle myosin regulation by light chain phosphorylation. The effects of phosphorylation of the regulatory light chains by either myosin light chain kinase or protein kinase C upon these distances will be examined. Using these approaches, the hypothesis that the head-tail junction of the myosin molecule plays a key role in the mechanism of regulation of smooth muscle myosin will be tested. This information will be useful to determine how the phosphorylation dependent conformational changes observed in vitro relate to physiologically important structural changes within myosin filaments in vivo.