Cloning and sequencing of cDNA for fish slow skeletal muscle myosin light chains

不同种类的鱼都有两种类型的骨骼肌，即普通(白色)和黑色(红色)肌肉。有人认为，普通肌肉对应于高等脊椎动物的快肌肉，而暗肌肉对应于慢肌肉。这两种骨骼肌肌球蛋白都由两条约200 kDa的重链和四条约20 kDa的轻链组成，与高等脊椎动物的情况一样。鱼类快速骨骼肌肌球蛋白有三种轻链，分为两类，一对由两个异源成分组成的碱性轻链(按相对分子质量由大到小依次为A1和A2)和一对5，5‘-二硫-双-2-硝基苯甲酸(DTNB)轻链。另一方面，众所周知，鱼慢肌肌球蛋白与快肌肌球蛋白在轻链组成上存在差异。以前的肌球蛋白一般只有两种轻链(D1和D2)，心肌肌球蛋白也是如此。然而，关于鱼类慢速骨骼肌肌球蛋白轻链的结构特征的信息根本不可用。本研究分别克隆、测序和鉴定了鲜鱼和马鱼慢骨骼肌D1和D2轻链的基因克隆。与在A1轻链中检测到的一样，D1轻链在N末端具有部分差异肽部分。这一结果有力地表明，慢速骨骼肌肌球蛋白中的D1轻链与快速骨骼肌肌球蛋白中的A1轻链具有相同的功能。D2轻链也有丝氨酸残基，与快速肌肉的DTNA轻链相同，表现为磷酸化。这项研究的结果是已知的关于鱼慢肌肌球蛋白轻链的完整一级结构的首次报道。

Various species of fish have two types of skeletal muscle, the ordinary (white) and dark (red) muscles. It has been suggested that ordinary muscle corresponds to fast muscle of higher vertebrates, while dark muscle corresponds to slow muscle. Both skeletal muscle myosins consist of two heavy chains of approximately 200kDa and four light chains of approximately 20kDa as in the case of higher-vertebrate counterparts. The fish fast skeletal muscle myosin has been shown to have three kinds of light chains, which are classified into two groups, one pair of alkali light chains consisting of two heterologous components (A1 and A2 in the order of decreasing molecular weight) and another pair of 5,5'-dithio-bis-2-nitrobenzoic acid (DTNB) light chains. On the other hand, it is well-known that fish slow muscle myosin differs from fast muscle myosin in light chain composition. The former myosin generally has only two kinds of light chain (D1 and D2) as does cardiac myosin. However, information on structural characterization of fish slow skeletal muscle myosin light chains is not available at all. This study described cloning, sequencing, and characterization of cDNA clones encoding D1 and D2 light chains of striped jack Caranx delicatissimus and horse mackerel Trachurus japonicus slow skeletal muscles, respectively. D1 light chain was found to have a part of the difference peptide portion in the N-terminal region, as was detected in A1 light chains. This result strongly suggests that D1 light chain in slow skeletal muscle myosin possess the same function as A1 light chain in fast skeletal muscle myosin. D2 light chain was also found to have serine residue, showing phosphorylation as was detected in DTNA light chain of fast muscle. The results in this study were the first known reports on the complete primary structure of fish slow muscle myosin light chains.