Self-Association of Rabbit Muscle Phosphofructokinase: Effects of Ligands

Lyndal K. Hesterberg, James C. Lee

Research output: Contribution to journalArticle

47 Scopus citations

Abstract

The effects of ligands on the self-association of rabbit muscle phosphofructokinase (PFK) were investigated by velocity sedimentation at pH 7.0 and 23 °C. The concentration dependence of the weight-average sedimentation coefficient was monitored in the presence of these ligands. The mode of association and equilibrium constants characterizing each association step were determined by theoretical fitting of the sedimentation data. The simplest mode of association for the PFK system is M↔M2↔M4↔M16. Ligands and temperature would perturb the various equilibrium constants without altering the mode of association. The apparent equilibrium constants for the formation of tetramer, K4app, are increased in the presence of 0.1 mM ATP and 1.0 mM fructose 6-phosphate. The value of the sedimentation coefficient for the tetramer, s4°, that would best fit the data is 12.4 S instead of 13.5 S determined in the absence of substrates, thus implying a structural change in the tetramer induced by substrates. Only an insignificant amount of dimer is present under the experimental conditions. The presence of activators, ADP or phosphate, enhances the formation of tetramers, and s4° assumes a value of 13.5 S. Similar results are obtained with decreasing concentrations of proton. The presence of the inhibitor, citrate, however, favors the formation of dimers. The equilibrium constants determined as a function of ADP concentration were further analyzed by the linked-function theory derived by Wyman [Wyman, J. (1964) Adv. Protein Chem. 19, 224-285], leading to the conclusion that the formation of a tetramer involves the binding of two additional molecules of ADP per monomer. Similar analysis results in a conclusion that the formation of a dimer involves the binding of one additional molecule of citrate per phosphofructokinase subunit.

Original languageEnglish (US)
Pages (from-to)216-222
Number of pages7
JournalBiochemistry
Volume21
Issue number2
DOIs
StatePublished - Jan 1982

ASJC Scopus subject areas

  • Biochemistry

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