TE-KINASE-GLYCERALDEHYDE-3-PHOSPHATE-DEHYDROGENASE INTERACTION - MOLECULAR-MASS STUDIES

When rabbit muscle phosphoglycerate kinase (PGK; a 48-kDa monomeric pr otein) and glyceraldehyde-3-phosphate dehydrogenase (GraPDH; a 145-kDa homotetrameric protein) are present together in solution in the propo rtion of 1 mol PGK/1 mol GraPDH monomer (total protein 0.2-1.0 mg/ml), an 80-82-kDa protein species is observed by gel-penetration (dilution factor) method and by the conventional procedure of elution from a ge l column. Individually, PGK and GraPDH do not exhibit any self associa tion or dissociation in the concentration range employed. Electrophore sis of the 80-82-kDa peak eluted from the gel column shows a single pr otein band with mobility intermediate between those of GraPDH and PGK. In titration experiments by the gel-penetration method, plots of dilu tion factor of PGK (or GraPDH) activity versus GraPDH (or PGK) concent ration shows two linear portions intersecting at approximately 1 mol G raPDH monomer/1 mol PGK. From the molecular-mass values and the titrat ion experiments, it has been suggested that, in solution, these enzyme s form a complex consisting of 1 molecule of PGK and one monomeric sub unit of GraPDH (expected molecular mass 84 kDa). Its dissociation cons tant has been estimated to be equal to or less than 13 nM. The complex is dissociated in the presence of KCl or NADH, with approximately hal f dissociation at 0.1 M salt or 0.25 mM NADH. At 0.1 M KCl, the comple x is completely dissociated by adding ATP, NADH or 3-phosphoglycerate. AMP, ADP, NAD(+), glyceraldehyde-3-phosphate, phosphate ions and fruc tose 1,6-bisphosphate reverse the effect of KCl.