Micellar electrokinetic chromatography as a complementary method to sodiumdodecyl sulfate-polyacrylamide gel electrophoresis for studying limited proteolysis of proteins

Micellar electrokinetic chromatography (MEKC) has been utilized as an analy tical method to perform investigations on limited proteolysis of proteins. To this purpose partial proteolysis experiments with a series of proteinase s were performed, utilizing as model protein pyruvate kinase (PK) from Esch erichia coil, an enzyme that is regulated allosterically by fructose 1,6-bi sphosphate (FBP). Data obtained with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and MEKC were compared; the profiles generat ed by submitting digests of PK treated with different proteinases in the pr esence and absence of FBP to electrophoretic analysis provided a useful adj unct for a better understanding of the effects or the allosteric activator on the conformation of the model enzyme. MEKC was also found to be a conven ient technique for determining the kinetics of substrate proteolysis.