pH-induced hysteretic properties of phosphofructokinase purified from rat myocardium.
Phosphofructokinase (PFK) purified from the rat myocardium is reversibly inactivated under a pH regime approximating that reported for ischemic hearts. At pH 6.5 and 37 degrees C, the enzyme displays a hysteretic loss of activity during 60-min incubations, declining to 48% of control (pH 7.1, 37 degrees C) values. Citric acid increases the degree of inactivation (28% of control), whereas fructose 1,6-bisphosphate reduces the decline in activity. Simultaneous measurements of 90 decreases light scattering and catalytic activity suggest the inactivation is temporally linked to dissociation of active tetrameric enzyme into an inactive form of lower molecular weight. Fluorescence enhancement of the extrinsic probe sodium mansate, which binds preferentially to dimeric PFK, indicates that the equilibrium dimer concentration (cp1 infinity) increases as pH is lowered. This increase in cp1 infinity exhibits a strong inverse correlation (r = 0.984) with catalytic activity across the pH range of 8.0 to 6.5. Returning solution pH to 7.0 or above promotes a time-dependent reactivation and repolymerization of PFK. The rate of reactivation is increased at higher enzyme concentrations and in the presence of trimethylamine-N-oxide, a nitrogenous osmolyte noted for its ability to promote protein aggregation reactions. Thus these results demonstrate the capacity of rat heart PFK to undergo reversible inactivation and dissociation in vitro and represent the first phase of a two-part study testing the hypothesis that these pH-induced hysteretic processes are operative in the ischemic myocardium. The data are evaluated in terms of the potential roles of hysteretic enzymes in metabolic regulation.
D000818:Animals; D003327:Coronary Disease / Q000201:enzymology / Q000503:physiopathology; D004789:Enzyme Activation; D005453:Fluorescence; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008027:Light; D008297:Male; D009206:Myocardium / Q000201:enzymology*; D009282:Naphthalenesulfonates; D010732:Phosphofructokinase-1 / Q000302:isolation & purification*; D011108:Polymers; D051381:Rats; D011919:Rats, Inbred Strains; D012542:Scattering, Radiation; D013997:Time Factors
D009282:Naphthalenesulfonates; D011108:Polymers; C009497:2-(4-toluidino)-6-naphthalenesulfonic acid; D010732:Phosphofructokinase-1