A simple procedure for the isolation of seven abundant muscle enzymes.
The present work describes procedures in which seven major muscle enzymes and serum albumin can be simultaneously isolated from chicken skeletal muscles. The seven enzymes isolated were: phosphorylase, enolase, creatine-P kinase, aldolase, glyceraldehyde-3-P dehydrogenase, phosphoglycerate mutase, and triose-P isomerase. The proteins isolated by these methods were judged to be greater than 97% pure on the basis of electrophoretic analysis in sodium dodecyl sulfate polyacrylamide gels. The procedure is applicable for isolation of the enzymes from large (greater than 100 g) or small (less than 0.5 g) amounts of muscle tissue and the entire procedure can be completed within two days. Particularly useful features of the procedures are: (1) preferential solubilization of the enzymes from myofibrils by extraction of muscle specimens in solutions of different ionic strength; (2) specific precipitation of phosphorylase, creatine-P kinase, and glyceraldehyde 3-Phosphate dehydrogenase from solutions of specified pH and degrees of ammonium sulfate saturation; and (3) an alternate method for isolation of glyceraldehyde-3-P dehydrogenase by specific elution of the enzyme from phosphocellulose columns with ATP. Because of the ease, rapidity, and reproducibility of the procedures, these methods may be useful for the routine isolation of the muscle enzymes in studies on biochemical regulation, as well as for obtaining large quantitites of the enzymes for structural analysis.
D000818:Animals; D002238:Carbohydrate Epimerases / Q000302:isolation & purification*; D002645:Chickens; D003402:Creatine Kinase / Q000302:isolation & purification*; D005634:Fructose-Bisphosphate Aldolase / Q000302:isolation & purification*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000302:isolation & purification*; D008722:Methods; D009132:Muscles / Q000201:enzymology*; D009210:Myofibrils / Q000201:enzymology; D009994:Osmolar Concentration; D010736:Phosphoglycerate Mutase / Q000302:isolation & purification*; D010751:Phosphopyruvate Hydratase / Q000302:isolation & purification*; D006005:Phosphorylases / Q000302:isolation & purification*; D010770:Phosphotransferases / Q000302:isolation & purification*; D014305:Triose-Phosphate Isomerase / Q000302:isolation & purification*
D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D006005:Phosphorylases; D010770:Phosphotransferases; D003402:Creatine Kinase; D005634:Fructose-Bisphosphate Aldolase; D010751:Phosphopyruvate Hydratase; D002238:Carbohydrate Epimerases; D014305:Triose-Phosphate Isomerase; D010736:Phosphoglycerate Mutase