Helfman D M, Feramisco J R, Ricci W M, Hughes S H
J Biol Chem. 1984 Nov 25;259(22):14136-43.
We have constructed a cDNA-expression library of approximately 100,000 members from embryonic chicken smooth-muscle mRNA using the plasmid-expression vectors pUC8 and pUC9. Using an immunological screening procedure and 32P-labeled cDNA probes, we have identified and isolated clones encoding smooth-muscle tropomyosin. Plasmid pSMT-10 (approximately 1100 base pairs) was found to hybrid-select mRNA for smooth-muscle alpha-tropomyosin. DNA-sequence analysis revealed that pSMT-10 contained the entire coding region for alpha-tropomyosin and portions of the 5'- and 3'-untranslated regions. Comparison of the derived amino acid sequence of smooth-muscle alpha-tropomyosin with known skeletal-muscle (rabbit and chicken) and platelet (equine) sequences revealed extensive homology between the various proteins. The smooth-muscle tropomyosin shows the greatest sequence divergence from the skeletal-muscle tropomyosins at the COOH-terminal region. In contrast, the smooth-muscle tropomyosin is most homologous to the platelet tropomyosin at the COOH-terminal end. The relationship of the various tropomyosin sequences to function (e.g. interactions with troponin) are considered.
我们使用质粒表达载体pUC8和pUC9，构建了一个由大约100,000个成员组成的来自胚胎鸡平滑肌mRNA的cDNA表达文库。通过免疫筛选程序和32P标记的cDNA探针，我们鉴定并分离出了编码平滑肌原肌球蛋白的克隆。发现质粒pSMT - 10（约1100个碱基对）能杂交选择平滑肌α - 原肌球蛋白的mRNA。DNA序列分析表明，pSMT - 10包含α - 原肌球蛋白的完整编码区以及5'和3'非翻译区的部分序列。将平滑肌α - 原肌球蛋白的推导氨基酸序列与已知的骨骼肌（兔和鸡）及血小板（马）序列进行比较，发现各种蛋白质之间存在广泛的同源性。平滑肌原肌球蛋白在COOH末端区域与骨骼肌原肌球蛋白的序列差异最大。相比之下，平滑肌原肌球蛋白在COOH末端与血小板原肌球蛋白最为同源。还考虑了各种原肌球蛋白序列与功能的关系（例如与肌钙蛋白的相互作用）。
