Yoshida M, Suzuki A, Yamamoto H, Noguchi S, Mizuno Y, Ozawa E
Department of Cell Biology, National Institute of Neuroscience, Tokyo, Japan.
Eur J Biochem. 1994 Jun 15;222(3):1055-61. doi: 10.1111/j.1432-1033.1994.tb18958.x.
Dystrophin is purified as a complex with several proteins from the digitonin-solubilized muscle cell membrane. Most of dystrophin-associated proteins (DAPs) are assumed to form a large oligomeric transmembranous glycoprotein complex on the sarcolemma and link dystrophin with a basement membrane protein, laminin. In the present study, we found that the purified dystrophin-DAP complex was dissociated into several groups by n-octyl-beta-D-glucoside treatment. In particular, we found that the glycoprotein complex stated above was dissociated into two distinct groups: one composed of 156DAG and 43DAG (A3a) and the other composed of 50DAG, 35DAG and A3b. We confirmed by crosslinking and immunoaffinity chromatography that these two groups existed in a complexes. We thus concluded that the glycoprotein complex consists of these two subcomplexes. Furthermore, A3b and 43DAG, which had been formerly treated simply as the 43DAG doublets due to their similar electrophoretic mobilities in SDS/PAGE, were shown to be present in two different subcomplexes. Based on the analyses by two-dimensional gel electrophoresis, peptide mapping and immunoblotting, we concluded that A3b is a novel DAP different from 43DAG.
肌营养不良蛋白与来自洋地黄皂苷增溶的肌细胞膜的几种蛋白质形成复合物后被纯化。大多数肌营养不良蛋白相关蛋白(DAPs)被认为在肌膜上形成一个大的寡聚跨膜糖蛋白复合物,并将肌营养不良蛋白与基底膜蛋白层粘连蛋白连接起来。在本研究中,我们发现纯化的肌营养不良蛋白 - DAP复合物经正辛基 - β - D - 葡萄糖苷处理后解离成几组。特别是,我们发现上述糖蛋白复合物解离成两个不同的组:一组由156DAG和43DAG(A3a)组成,另一组由50DAG、35DAG和A3b组成。我们通过交联和免疫亲和层析证实这两组存在于复合物中。因此我们得出结论,糖蛋白复合物由这两个亚复合物组成。此外,A3b和43DAG,由于它们在SDS/PAGE中相似的电泳迁移率,以前被简单地视为43DAG双峰,现在显示它们存在于两个不同的亚复合物中。基于二维凝胶电泳、肽图谱分析和免疫印迹分析,我们得出结论,A3b是一种不同于43DAG的新型DAP。