Quantitative determination of Na+-K+-ATPase and other sarcolemmal components in muscle cells.
A recurring problem in the characterization of plasma membrane enzymes in tissues and cells is whether the samples tested are representative for the entire population of enzyme molecules present in the starting material. Measurements of [3H]-ouabain binding, enzyme activity, and maximum transport capacity all indicate that the concentration of Na+-K+ pumps in mammalian skeletal muscle is high (300-800 pmol/g wet wt). Studies on Na+-K+-ATPase activity in isolated sarcolemma, however, generally give little or no information on total cellular enzyme concentration. Due to the low and variable enzyme recovery (0.2-8.9%), such subcellular preparations may, therefore, give misleading data on factors regulating Na+-K+-ATPase in heart and skeletal muscle cells. As the same isolation and purification procedures are used for the study of other sarcolemmal components (lipids, hormone receptors, enzymes, and other transport systems), this inadequate recovery has general implications for statements on regulatory changes in the sarcolemmal composition of muscle cells. On the other hand, complete quantification of Na+-K+-ATPase in muscle tissue can now be achieved using simple procedures and the entire material (intact muscle fibers, biopsies, and whole homogenates). Recent studies have shown that regulatory changes in the entire population of Na+-K+ pumps in muscle can be quantified in measurements of [3H]-ouabain binding, K+-activated 3-O-methylfluorescein phosphatase activity, as well as maximum ouabain suppressible Na+-K+ transport capacity.
D000818:Animals; D001693:Biological Transport, Active; D007473:Ion Channels / Q000378:metabolism; D009132:Muscles / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism; D009994:Osmolar Concentration; D010042:Ouabain / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D011188:Potassium / Q000502:physiology; D012508:Sarcolemma / Q000201:enzymology* / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D007473:Ion Channels; D010042:Ouabain; D012964:Sodium; D010744:Phosphoric Monoester Hydrolases; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium