Guerriero V, Rowley D R, Means A R
Department of Cell Biology, Baylor College of Medicine, Houston, Texas 77030.
Cell. 1981 Dec;27(3 Pt 2):449-58. doi: 10.1016/0092-8674(81)90386-x.
A specific precipitating antibody against chicken gizzard myosin light chain kinase (MLCK) has been produced in rabbits. The antibody inhibited enzyme activity in a dose-dependent manner with 11 moles of antibody required to inhibit 80% of the activity of one mole MLCK. Crude homogenates from various chicken tissues were analyzed by SDS-polyacrylamide gel electrophoresis, and the proteins were transferred onto nitrocellulose sheets and reacted with antibody. In each case, the antibody bound to only one protein with a molecular weight of approximately 130,000. These data suggest the MLCK present in all types of muscle as well as non-muscle tissues is of the same molecular weight. Indirect immunofluorescent microscopy of non-muscle tissue culture cells revealed MLCK to be localized in the spindle apparatus and midbody of mitotic cells and on the stress fibers and in the nucleolus of interphase cells. The nucleolar localization was confirmed by electron microscopy and shown to be restricted to the fibrillar region. In myofibrils isolated from skeletal and cardiac muscle, anti-MLCK decorated the actin-containing I bands of the sarcomere. These results are consistent with the suggestion that MLCK and calmodulin are intermediates in the regulation of cell motility by calcium.
已在兔体内产生了一种针对鸡砂囊肌球蛋白轻链激酶（MLCK）的特异性沉淀抗体。该抗体以剂量依赖方式抑制酶活性，抑制一摩尔MLCK活性的80%需要11摩尔抗体。通过SDS-聚丙烯酰胺凝胶电泳分析来自各种鸡组织的粗匀浆，然后将蛋白质转移到硝酸纤维素膜上并与抗体反应。在每种情况下，抗体仅与一种分子量约为130,000的蛋白质结合。这些数据表明，存在于所有类型肌肉以及非肌肉组织中的MLCK具有相同的分子量。对非肌肉组织培养细胞进行间接免疫荧光显微镜检查发现，MLCK定位于有丝分裂细胞的纺锤体装置和中间体以及间期细胞的应力纤维和核仁中。核仁定位通过电子显微镜得到证实，并显示局限于纤维状区域。在从骨骼肌和心肌分离的肌原纤维中，抗MLCK抗体标记了肌节中含肌动蛋白的I带。这些结果与以下观点一致，即MLCK和钙调蛋白是钙调节细胞运动的中间介质。
