Rat liver phosphofructokinase: kinetic and physiological ramifications of the aggregation behavior. Academic Article uri icon


  • The effect of allosteric ligands on the aggregation of rat liver phosphofructokinase (PFK) was examined by measuring the fluorescence polarization of pyrenebutyratephosphofructokinase (PB-PFK) as described in the preceding paper. At a protein concentration of 8 jug/mL, near-saturating concentrations of fructose 1, 6-bisphosphate, phosphate, or ammonium sulfate stabilize fairly high degrees of polarization whereas AMP and citrate promote polarization values comparable to or below that in the presence of MgATP. However, when each ligand is present in combination with either MgATP or fructose 6-phosphate (F6P), the resulting polarization is very similar to that observed with MgATP or F6P alone, indicating that the influence of MgATP or F6P dominates that of the other ligands. the effect of magnesium adenylyl imidodiphosphate (MgAMPPNP) on the polarization of PB-PFK is very similar to that of MgATP. In the presence of MgAMPPNP and increasing amounts of F6P, the polarization increases, indicating that as F6P binds the aggregation equilibria are shifted toward association. In the presence of MgAMPPNP and low levels of F6P, activators that promote F6P binding also promote aggregation. Activators also promote F6P binding to a small enzyme population with subsequent aggregation and reactivation. These observations are supportive of the hypothesis that F6P binds more favorably to the high MT aggregate population than it does to the tetramer or smaller populations. Gel filtration of lOOOOOg X 60 min supernatant fractions of rat liver homogenates, as well as the ligand effects described, supports the conclusion that a substantial portion of the PFK in the cell exists as a high Mr aggregate. Hence, it is proposed that PFK in vivo binds F6P better than during the highly diluted conditions of kinetic assays in vitro and should therefore be more active at the low F6P concentration found in the cell. This proposal is supported by the finding that specific activity increases with PFK concentration when measured under inhibiting conditions. © 1980, American Chemical Society. All rights reserved.

published proceedings

  • Biochemistry

author list (cited authors)

  • Reinhart, G. D., & Lardy, H. A

citation count

  • 46

complete list of authors

  • Reinhart, GD||Lardy, HA

publication date

  • April 1980