Purification and properties of mouse pyruvate kinases K and M and of a modified K subunit

Abstract

The K4 and M4 isozymes of mouse pyruvate kinase were purified to homogeneity, and their physical, chemical, and kinetic properties were compared. The K isozyme is slightly larger, but a high degree of homology exists as evidenced by a similar amino acid composition, immunotitration value, and two-dimensional arginine peptide pattern after tryptic digestion. Also, the more active conformational form of the K isozyme has kinetic and chromatographic properties similar to those of the M isozyme. Only K subunit could be extracted with antibody from fresh spleen extracts, but this subunit can be cleaved to form a product with the mobility of the M subunit. The cleavage is accomplished by an endogenous enzyme and appears to be the first step in K-enzyme degradation. This product is called Kpm . KpmK hybrid could also be purified to homogeneity. This enzyme has the structure K2pmK2, and both types of subunit have activity. The Kpm form has a higher K0.5S value for phosphoenolpyruvate and a lower K0.5S value for ADP than does either the K or the M type. However, the Kpm and M subunits otherwise have very similar properties and it is speculated that the Kpm subunit is an M-type precursor.