Interaction of the Dissociable Glycerol‐3‐phosphate Dehydrogenase and Fructose‐1,6‐bisphosphate Aldolase

Cytoplasmic sn ‐glycerol‐3‐phosphate dehydrogenase, labelled covalently with fluorescein isothiocyanate, shows an enzyme‐concentration‐dependent fluorescence anisotropy. The anisotropy versus enzyme concentration curves is shifted towards higher concentrations when substrates are present. The comparison of the dissociation constants estimated from anisotropy measurements and derived from kinetic experiments suggests that the substrate‐induced dissociation of the dimeric dehydrogenase is slow with respect to the enzymatic reaction catalyzed by either its monomeric or dimeric form. The fluorescence anisotropy of the fluorescent dye‐labelled dehydrogenase increases with time upon addition of unlabelled fructose‐1,6‐bisphosphate aldolase approaching a limiting value. This fact indicates the binding of fructose‐1,6‐bisphosphate aldolase to glycerolphosphate dehydrogenase. A model is proposed assuming simuitaneous binding of tetrameric fructose‐1,6‐bisphosphate aldolase to monomeric and dimeric glycerolphosphate dehydrogenase with 1:1 stoichiometry. The dissociation constants, as parameters fitted to the experimental curves, were estimated as 0.2 μM and 1 μM for aldolase–dimeric‐glycerolphosphate‐dehydrogenase and aldolase–monomeric‐glycerolphosphate‐dehydrogenase complexes respectively.