Purification of glycolytic enzymes by using affinity-elution chromatography.
1. A systematic procedure for the purification of enzymes by affinity-elution chromatography is described. Enzymes are adsorbed on a cation-exchanger, and eluted with ligands specific for the enzyme concerned. 2. All of the glycolytic and some related enzymes present in rabbit muscle can be purified by the affinity-elution technique. The pH range for adsorption and elution of each enzyme was found, and the effects of minor variations of conditions are described. 3. A description of experimental conditions suitable for affinity elution of each enzyme is given, together with special features relevant to each individual enzyme. 4. Theoretical considerations of affinity elution chromatography are discussed, including its limitations, advantages and disadvantages compared with affinity-adsorption chromatography. Possible developments are suggested to cover enzymes which because of their adsorption characteristics are not at present amenable to affinity-elution procedures.
D000263:Adenylate Kinase / Q000302:isolation & purification; D000818:Animals; D002846:Chromatography, Affinity / Q000379:methods*; D003402:Creatine Kinase / Q000302:isolation & purification; D004798:Enzymes / Q000302:isolation & purification*; D005634:Fructose-Bisphosphate Aldolase / Q000302:isolation & purification; D005956:Glucose-6-Phosphate Isomerase / Q000302:isolation & purification; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000302:isolation & purification; D006019:Glycolysis; D007770:L-Lactate Dehydrogenase / Q000302:isolation & purification; D009132:Muscles / Q000201:enzymology; D010732:Phosphofructokinase-1 / Q000302:isolation & purification; D010733:Phosphoglucomutase / Q000302:isolation & purification; D010735:Phosphoglycerate Kinase / Q000302:isolation & purification; D010736:Phosphoglycerate Mutase / Q000302:isolation & purification; D010751:Phosphopyruvate Hydratase / Q000302:isolation & purification; D006005:Phosphorylases / Q000302:isolation & purification; D011770:Pyruvate Kinase / Q000302:isolation & purification; D011817:Rabbits; D014305:Triose-Phosphate Isomerase / Q000302:isolation & purification
D004798:Enzymes; D007770:L-Lactate Dehydrogenase; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D006005:Phosphorylases; D010732:Phosphofructokinase-1; D011770:Pyruvate Kinase; D010735:Phosphoglycerate Kinase; D003402:Creatine Kinase; D000263:Adenylate Kinase; D005634:Fructose-Bisphosphate Aldolase; D010751:Phosphopyruvate Hydratase; D014305:Triose-Phosphate Isomerase; D005956:Glucose-6-Phosphate Isomerase; D010736:Phosphoglycerate Mutase; D010733:Phosphoglucomutase