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首页> 外文期刊>Molecular and Cellular Biology >Dictyostelium discoideum myosin: isolation and characterization of cDNAs encoding the regulatory light chain.
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Dictyostelium discoideum myosin: isolation and characterization of cDNAs encoding the regulatory light chain.

机译：盘基网柄菌肌球蛋白：编码调节性轻链的cDNA的分离和鉴定。

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Phosphorylation of the regulatory light chains (RMLC) of nonmuscle myosin can increase the actin-activated ATPase activity and filament formation. Little is known about these regulatory mechanisms and how the RMLC are involved in ATP hydrolysis. To better characterize the nonmuscle RMLC, we isolated cDNAs encoding the Dictyostelium RMLC. Using an antibody specific for the RMLC, we screened a lambda gt11 expression library and obtained a 200-base-pair clone that encoded a portion of the RMLC. The remainder of the sequence was obtained from two clones identified by DNA hybridization, using the 200-base-pair cDNA. The composite RMLC cDNA was 645 nucleotides long. It contained 60 base pairs of 5' untranslated, 483 bases of coding, and 102 base pairs of 3' untranslated sequence. The amino acid sequence predicted an 18,300-dalton protein that shares 42% amino acid identity with Dictyostelium calmodulin and 30% identity with the chicken skeletal myosin RMLC. This sequence contained three regions that were similar to the E-F hand calcium-binding domains found in calmodulin, troponin C, and other myosin light chains. A sequence similar to the phosphorylation sequence found in chicken gizzard and skeletal myosin light chains was found at the amino terminus. Genomic Southern blot analysis suggested that the Dictyostelium genome contains a single gene encoding the RMLC. Analysis of RMLC expression patterns during Dictyostelium development indicated that accumulation of this mRNA increases just before aggregation and again during culmination. This pattern is similar to that obtained for the Dictyostelium essential myosin light chain and suggests that expression of the two light chains is coordinated during development.
机译：非肌肉肌球蛋白的调节性轻链（RMLC）的磷酸化可以增加肌动蛋白激活的ATPase活性和细丝形成。关于这些调节机制以及RMLC如何参与ATP水解知之甚少。为了更好地表征非肌肉RMLC，我们分离了编码小球藻RMLC的cDNA。使用针对RMLC的特异性抗体，我们筛选了λgt11表达文库，并获得了200个碱基对的克隆，该克隆编码了RMLC的一部分。该序列的其余部分是使用200个碱基对的cDNA从通过DNA杂交鉴定的两个克隆中获得的。复合RMLC cDNA长645个核苷酸。它包含60个5'非翻译的碱基对，483个编码碱基和102个3'非翻译序列的碱基对。该氨基酸序列预测了一种18,300道尔顿的蛋白质，与Dictyostelium钙调蛋白具有42％的氨基酸同一性，与鸡骨骼肌肌球蛋白RMLC具有30％的同一性。该序列包含三个区域，它们与钙调蛋白，肌钙蛋白C和其他肌球蛋白轻链中的E-F手钙结合结构域相似。在氨基末端发现了类似于在鸡g和骨骼肌肌球蛋白轻链中发现的磷酸化序列的序列。基因组Southern印迹分析表明，网盘菌属基因组包含编码RMLC的单个基因。在盘基网柄菌发育期间RMLC表达模式的分析表明，该mRNA的积累在聚集之前以及在高潮期间再次增加。这种模式类似于从网柄菌属必需的肌球蛋白轻链获得的模式，表明在发育过程中两个轻链的表达是协调的。

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