BIOCHEMISTRY OF CONTRACTILE PROTEINS

我们的长期目标是更好地了解监管 平滑肌中的机制。 平滑肌对正常的身体至关重要 功能，是动脉，静脉，肠， 子宫等。众所周知，细胞内 高浓度的CA++诱导平滑肌收缩。 机制 CA++浓度的变化通过收缩细胞检测， 然后进行处理，以导致收缩或松弛 仍然存在争议。 有两种基本理论：最流行的 假设是肌球蛋白被肌球蛋白轻链激酶磷酸化 （MLCK）激活收缩器官并导致收缩， 失活（松弛）是通过肌球蛋白的去磷酸化来实现的， 肌球蛋白轻链磷酸酶（MLCP）;另一种理论是， 调节是通过一种称为leitonin的机制实现的， 肌球蛋白磷酸化 有大量证据表明， 磷酸化理论至少构成了调节机制的一部分 这一建议的重点是详细审查的不同方面， 这个理论。 MLCK与收缩因子的相关性 将被研究。 有证据表明MLCK与肌动蛋白结合。 这将是 确认并建立约束条件。 如果MLCK是 仅限于细丝，细丝重叠的程度将决定 磷酸化水平。 改进的隔离程序 将对MLCK进行调查。 有一些数据表明， 先前分离的MLCK可以源自较大的前体。 的 磷酸化反应也将更详细地分析，因为它是 磷酸化可能是连续的而不是随机的， 这是肌球蛋白分子的一般性质，将通过研究 平滑肌和骨骼肌肌球蛋白的磷酸化， 它们的亚片段 一个重要的目标是确立 磷酸化和几个实验建议，其中 磷酸化可与肌动球蛋白的ATP酶活性相关， 超降水 是否非循环肌球蛋白-肌动蛋白附着 将研究在体外条件下形成的。 的数据 将对上述实验进行评价，以评估肌球蛋白的作用。 磷酸化，并质疑是否有其他机制 指出了 后者也将使用CA++不敏感MLCK进行测试。

Our long range goal is to gain a better understanding of the regulatory mechanisms in smooth muscle. Smooth muscle is essential for normal body function and is the contractile element of arteries, veins, intestines, uterus, etc. It is known that an increase in the intracellular concentration of CA++ induces contraction of smooth muscle. The mechanism by which the changes in CA++ concentration are detected by the contractile apparatus and then processed to result in either contraction or relaxation is still controversial. There are two basic theories: the most popular hypothesis is that phosphorylation of myosin by a myosin light chain kinase (MLCK) activates the contractile apparatus and leads to contraction, deactivation (relaxation) is achieved by dephosphorylation of myosin by a myosin light chain phophatase (MLCP); the alternative theory is that regulation is achieved by a mechanism termed leiotonin and does not involve myosin phosphorylation. There is considerable evidence to indicate that the phosphorylation theory forms at least part of the regulatory mechanism and the focus of this proposal is to examine in detail different aspects of this theory. The association of the MLCK with the contractile elements will be studied. Evidence suggests that MLCK binds to actin. This will be confirmed and the conditions of binding established. If the MLCK is restricted to thin filaments the extent of filament overlap will dictate the level of phosphorylation. Improved procedures for the isolation of MLCK will be investigated. There are some data to sugget that all previously isolated MLCKs may be derived from a larger precursor. The phosphorylation reaction will also be analyzed in more detail since it is possible that phosphorylation is sequential rather than random, and whether this is a general property of myosin molecules will be tested by studying phosphorylation of smooth muscle and skeletal muscle muscle myosins and their subfragments. An important objective is to estgablish the role of phosphorylation and several experiments are suggested in which phosphorylation can be correlated to ATPase activity of actomyosin or superprecipitation. Whether or not non-cycling myosin-actin attachments are formed under in vitro conditions will be investigated. The data from the above experiments will be evaluated to assess the role of myosin phosphorylation and to question whether additional mechanisms are indicated. The latter will also be tested using CA++ insensitive MLCK.