BIOCHEMISTRY AND BIOLOGY OF A MYOSIN BINDING PROTEIN

非肌肉细胞调节重要的ACTO的方式。 肌球蛋白介导的形状变化负责细胞分裂， 伪足伸展、内吞和许多其他形式的 运动还没有完全被理解。学识渊博 存在关于各种肌动蛋白结合的生物化学和生物学 调节细胞肌动蛋白组装的蛋白质。另一方面 手，虽然对肌球蛋白的生物化学知道很多 通过磷酸化进行调节，人们对细胞如何 改变肌球蛋白在细胞中的组装状态和定位 有动静。肌肉细胞有许多与肌球蛋白相关的 包括轻链和重链激酶在内的蛋白质，它们介导 肌球蛋白组装，以及星云蛋白、肌蛋白和C蛋白的一部分 粗大的细丝。然而，人们对非肌肉组织知之甚少。 细胞肌球蛋白相关蛋白，而不是 激活剂。有趣的是，肌球蛋白表现出非常低的离子 在高速细胞提取液、条件下的强度溶解度 在那里，当纯净时，它会组装成两极细丝，可以凝胶化 与肌动蛋白及其相关蛋白结合，并在肌球蛋白中收缩- 依赖的态度。我们发现了一种53kD的新蛋白质 亚基分子量为海胆卵提取液 直接调解这一通常的低离子强度溶解度误差 肌球蛋白通过结合影响肌球蛋白低离子强度的溶解度 直接以肌球蛋白影响肌球蛋白的低离子强度溶解度。 我们也有初步证据表明53kD结合在头部- 肌球蛋白的杆状连接并表现出钙调素非依赖性 肌球蛋白轻链激酶活性。在这项提案中，我们打算 进一步描述这种蛋白质的特性：与 其他轻链激酶、功能结构域、准确结合部位 肌球蛋白，磷酸化调节，对肌球蛋白的影响 ATPase和组装，以及无处不在。一系列生物学上的 将通过使用in来询问有关53 kD函数的问题 体外肌球蛋白介导的运动性测定 肌球蛋白在运动性上被53kD结合或磷酸化。 53kD在体内的作用将通过分析其影响来研究 微量注射抗53kD抑制性单抗的研究 海胆受精卵的卵裂活性及其定位 在这个细胞和其他细胞中，53kD在整个细胞周期中。这些 对一种新蛋白质的研究应该有助于弥合我们之间的差距 肌球蛋白活性调控的分子知识及其调控方式 该细胞控制肌球蛋白的动态组装和定位 有动静。

The way in which non-muscle cells regulate the important acto- myosin mediated shape changes responsible for cell division, pseudopod extension, endocytosis, and many other forms of movement are not fully understood. A great deal of knowledge exists about the biochemistry and biology of various actin binding proteins which regulate cellular actin assemblies. On the other hand, while a good deal is known about the biochemistry of myosin regulation by phosphorylation, little is known about how the cell changes the assembly state and localization of myosin during cell movement. Muscle cells have a number of myosin-associated proteins including light and heavy chain kinases, which mediate myosin assembly, and nebulin, titin, and C-protein which are part of the thick filament. However, little is known about non-muscle cell myosin-associated proteins other than the biochemistry of the kinases. Interestingly, myosin exhibits very unusual low ionic strength solubility in high speed cellular extracts, conditions where when pure it assembles into bipolar filament, which can gel with actin and its associated proteins and contract in a myosin- dependent manner. We have identified a new protein of 53 kD subunit molecular weight is sea urchin egg extracts which mediates this ususual low ionic strength solubility error directly to myosin to affect myosin low ionic strength solubility by binding directly to myosin to affect myosin low ionic strength solubility. We also have preliminary evidence that 53 kD binds to the head- rod junction of myosin and exhibits calmodulin-independent myosin light chain kinase activity. In this proposal we intend to further characterize this protein in terms of its: relatedness to other light chain kinases, functional domains, exact binding site on myosin, regulation by phosphorylation, effects on myosin ATPase and assembly, and ubiquity. A series of biological questions will be asked about 53 kD function through the use of in vitro myosin-mediated motility assays by studying the effects of myosin being bound by, or phosphorylated by, 53 kD on motility. The role of 53 kD in vivo will be studied by analyzing the effects of microinjected inhibitory monoclonal antibodies to 53 kD activity on cleavage of fertilized sea urchin eggs and by localizing 53 kD throughout the cell cycle in this and in other cells. These studies on a novel protein should help bridge the gap between our molecular knowledge of the regulation of myosin activity and how the cell controls myosin's dynamic assembly and localization for movement.