| Abstrakt: |
Three isoenzymes of yeast phosphoglucose isomerase were studied by physical and chemical methods to establish their subunit properties and to identify the structural differences among them which give rise to their heterogeneous chromatographic behavior. Equilibrium sedimentation ultracentrifugation in 6 mguanidine hydrochloride and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate showed all three isoenzymes to be dimers composed of subunits of equal size. (The latter technique was found to give rise to multiple band artifacts if special precautions were not taken to avoid interference from possible protease contaminations.) The subunit molecular weights of each isoenzyme species were determined to be 60,000, i.e. one-half of the molecular weight of 120,000 of the native, undissociated isoenzymes (Kempe, T. D., Nakagawa, Y., andNoltmann, E. A. (1974) J. Biol. Chem. 249, 4617–4624). Equilibrium binding studies with the competitive inhibitor pyridoxal 5′-phosphate indicated preferential binding at a single site per monomer and yielded a dissociation constant of 0.4 mm, in reasonable agreement with a kinetically derived Kiof 0.22 mm. |
