Samarel A M, Ferguson A G, Burgess L
Department of Medicine, Northwestern University Medical School, Chicago, IL 60611.
J Mol Cell Cardiol. 1987 Jul;19(7):699-707. doi: 10.1016/s0022-2828(87)80378-4.
We describe a method for the complete solubilization and quantitative analysis of individual myofibrillar proteins in whole tissue homogenates of ventricular myocardium using gradient dodecyl sulfate polyacrylamide gel electrophoresis and staining with 125I-labeled Coomassie brilliant blue. The procedure allows for the simultaneous quantification of myosin heavy chain, myosin light chain, phosphorylatable myosin light chain and actin from as little as 50 mg of tissue. Within-assay and between-assay variations range from 8.0% to 12.6% for each protein subunit. The method was applied to the determination of the subunit stoichiometry of purified myosin, and to the measurement of myosin and actin concentrations in the neonatal and adult rabbit heart. Furthermore, we provide quantitative biochemical evidence for the existence of large molar excesses of myosin light chains in tissue homogenates of both neonatal and adult rabbit ventricular myocardium.
我们描述了一种方法，用于使用梯度十二烷基硫酸钠聚丙烯酰胺凝胶电泳和用¹²⁵I标记的考马斯亮蓝染色，对心室心肌全组织匀浆中的单个肌原纤维蛋白进行完全溶解和定量分析。该方法允许从低至50毫克的组织中同时定量肌球蛋白重链、肌球蛋白轻链、可磷酸化肌球蛋白轻链和肌动蛋白。每种蛋白质亚基的测定内和测定间变异范围为8.0%至12.6%。该方法应用于纯化肌球蛋白亚基化学计量的测定，以及新生和成年兔心脏中肌球蛋白和肌动蛋白浓度的测量。此外，我们提供了定量生化证据，证明新生和成年兔心室心肌组织匀浆中存在大量摩尔过量的肌球蛋白轻链。
