Matusovsky Oleg S, Shelud'ko Nikolay S, Permyakova Tatyana V, Zukowska Magdalena, Sobieszek Apolinary
Institute for Biomedical Aging Research, Life Science Center, Austrian Academy of Sciences, Innsbruck, Austria.
Biochim Biophys Acta. 2010 Apr;1804(4):884-90. doi: 10.1016/j.bbapap.2009.12.020. Epub 2010 Jan 13.
We have shown previously that myorod, a molluscan thick filament protein of unknown function, is phosphorylated by vertebrate smooth myosin light chain kinase (MLCK) in N-terminal unique region. The aim of the present study was to clarify whether such phosphorylation may occur in molluscan muscles. We detected three kinases endogenous to molluscan catch muscle, namely, to the complex of surface thick filament proteins that consists of twitchin, myosin, and myorod. The first kinase was a protein kinase A because it was inhibited by a specific inhibitor; the second one was associated with twitchin and phosphorylated myorod at its N-terminal unique region independently of Ca(2+); and the third kinase was bound to myosin and phosphorylated myorod as well as myosin in the C-terminal part of both proteins. The myosin-associated kinase was inhibited by micromolar concentration of calcium ions. This enzyme could be separated from myosin by chromatography, whereas the kinase associated with twitchin could not be separated from twitchin. Since twitchin has a MLCK-like domain, it is possible that this domain was responsible for myorod phosphorylation. Phosphorylation of myorod within the twitchin-myosin-myorod complex increased the actin-activated Mg(2+)-ATPase activity of myosin. Taken together, these results indicate that phosphorylation of myorod by kinases associated with key proteins of catch contraction may contribute to the functional activity of myorod in molluscan smooth muscle.
我们之前已经表明，肌杆蛋白是一种功能未知的软体动物粗肌丝蛋白，在其N端独特区域被脊椎动物平滑肌肌球蛋白轻链激酶（MLCK）磷酸化。本研究的目的是阐明这种磷酸化是否会在软体动物肌肉中发生。我们检测到软体动物捕获肌内源性的三种激酶，即存在于由肌动蛋白结合蛋白、肌球蛋白和肌杆蛋白组成的表面粗肌丝蛋白复合物中。第一种激酶是蛋白激酶A，因为它被一种特异性抑制剂抑制；第二种激酶与肌动蛋白结合蛋白相关，并且在其N端独特区域独立于Ca(2+)使肌杆蛋白磷酸化；第三种激酶与肌球蛋白结合，并使肌杆蛋白以及这两种蛋白质C端部分的肌球蛋白磷酸化。与肌球蛋白相关的激酶被微摩尔浓度的钙离子抑制。这种酶可以通过色谱法与肌球蛋白分离，而与肌动蛋白结合蛋白相关的激酶不能与肌动蛋白结合蛋白分离。由于肌动蛋白结合蛋白有一个类似MLCK的结构域，所以这个结构域可能是肌杆蛋白磷酸化的原因。在肌动蛋白结合蛋白 - 肌球蛋白 - 肌杆蛋白复合物中肌杆蛋白的磷酸化增加了肌球蛋白的肌动蛋白激活的Mg(2+)-ATP酶活性。综上所述，这些结果表明，与捕获收缩关键蛋白相关的激酶对肌杆蛋白的磷酸化可能有助于肌杆蛋白在软体动物平滑肌中的功能活性。
