Phosphoglycerate-kinase-glyceraldehyde-3-phosphate-dehydrogenase interaction. Molecular mass studies.
When rabbit muscle phosphoglycerate kinase (PGK; a 48-kDa monomeric protein) and glyceraldehyde-3-phosphate dehydrogenase (GraPDH; a 145-kDa homotetrameric protein) are present together in solution in the proportion 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 gel column. Individually, PGK and GraPDH do not exhibit any self association or dissociation in the concentration range employed. Electrophoresis of the 80-82-kDa peak eluted from the gel column shows a single protein band with mobility intermediate between those of GraPDH and PGK. In titration experiments by the gel-penetration method, plots of dilution factor of PGK (or GraPDH) activity versus GraPDH (or PGK) concentration shows two linear portions intersecting at approximately 1 mol GraPDH monomer/1 mol PGK. From the molecular-mass values and the titration experiments, it has been suggested that, in solution, these enzymes form a complex consisting of 1 molecule of PGK and one monomeric subunit of GraPDH (expected molecular mass 84 kDa). Its dissociation constant 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 half dissociation at 0.1 M salt or 0.25 mM NADH. At 0.1 M KCl, the complex is completely dissociated by adding ATP, NADH or 3-phosphoglycerate. AMP, ADP, NAD+, glyceraldehyde-3-phosphate, phosphate ions and fructose-1,6-bisphosphate reverse the effect of KCl.
Department of Chemistry, Banaras Hindu University, Varanasi, India.
O P
OP
Malhotra
P
P
Prabhakar
T
T
Sen Gupta
A M
AM
Kayastha
D000818:Animals; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D008970:Molecular Weight; D009243:NAD; D010735:Phosphoglycerate Kinase / Q000378:metabolism*; D011189:Potassium Chloride; D011817:Rabbits
D009243:NAD; D011189:Potassium Chloride; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010735:Phosphoglycerate Kinase