Usefulness of the Linkage Concept to Understanding the Regulation of Rat Liver Phosphofructokinase

  • Gregory D. Reinhart
Conference paper


Although this tribute to Gregorio Weber has quite appropriately focused on the use of fluorescence methodologies in the study of macro-molecules, most of which he pioneered, it is perhaps also fitting to discuss other contributions to our current thinking regarding protein biophysics, that do not necessarily involve the exploitation of the fluorescence phenomenon, to which the Professor has made seminal contributions as well. I refer specifically to several papers (Weber, 1971, 1972, 1975), written in the early 1970’s, pertaining to protein ligand binding and the ramifications of the principles of linkage that must be satisfied when evaluating binding behavior. After a brief review of the points made in these papers, I would like to then discuss how the concept of linkage, as elaborated by Dr. Weber, has proven to be invaluable in providing a greater insight into the mechanisms governing the allosteric regulation of an enzyme of considerable interest to those studying the metabolic control of hepatic carbohydrate metabolism; namely, rat liver phosphofructokinase (PFK).


Free Energy American Chemical Society Fluorescence Polarization Standard Free Energy Rabbit Muscle 
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  1. Aaronson, R. P., and Frieden, C., 1971, Rabbit Muscle Phosphofructokinase: Studies on the Polymerization, J. Biol. Chem., 247:7502.Google Scholar
  2. Donofrio, J. C., Thompson, R. S., Reinhart, G. D., and Veneziale, C. M., 1984, Quantification of Liver and Kidney Phosphofructokinase by Radio-immunoassay in Fed., Starved, and Alloxan-Diabetic Rats, Biochem. J., 224:541.PubMedGoogle Scholar
  3. Furuya, E., and Uyeda, K., 1980a, An Activation Factor of Liver Phosphofructokinase, Proc. Natl. Acad. Sci. U.S.A., 77:5861.PubMedCrossRefGoogle Scholar
  4. Furuya, E., and Uyeda, K., 1980b, Regulation of Phosphofructokinase by a New Mechanism, J. Biol. Chem., 255:11656.PubMedGoogle Scholar
  5. Hers, H. G., and Van Schaftingen, E., 1982, Fructose 2,6-Bisphosphate 2 Years After Its Discovery, Biochem. J., 206:1.PubMedGoogle Scholar
  6. Hofer, H. W., 1971, Influence of Enzyme Concentration on the Kinetic Behavior of Rabbit Muscle Phosphofructokinase, Hoppe-Seylers Z. Physiol. Chem., 352:997.PubMedCrossRefGoogle Scholar
  7. Hulme, E. C., and Tipton, K. F., 1971, The Dependence of Phosphofructokinase Kinetics upon Protein Concentration, FEBS Lett., 12:197.PubMedCrossRefGoogle Scholar
  8. Knopp, J. A., and Weber, G., 1969, Fluorescence Polarization of Pyrene-butyric — Bovine Serum Albumin and Pyrenebutyric — Human Macroglobulin Conjugates, J. Biol. Chem., 244:6309.PubMedGoogle Scholar
  9. Lad, P. M., Hill, D. E., and Hammes, G. G., 1973, Influence of Allosteric Ligands on the Activity and Aggregation of Rabbit Muscle Phosphofructokinase, Biochemistry, 12:4303.PubMedCrossRefGoogle Scholar
  10. Pavelich, M. J., and Hammes, G. G., 1973, Aggregation of Rabbit Muscle Phosphofructokinase, Biochemistry, 12:1408.PubMedCrossRefGoogle Scholar
  11. Pettigrew, D. W., and Frieden, C., 1979a, Binding of Regulatory Ligands to Rabbit Muscle Phosphofructokinase. A Model for Nucleotide Binding as a Function of Temperature and pH, J. Biol. Chem., 254:1887.PubMedGoogle Scholar
  12. Pettigrew, D. W., and Frieden, C., 1979b, Rabbit Muscle Phosphofructokinase. A Model for Regulatory Kinetic Behavior, J. Biol. Chem., 254:1896.PubMedGoogle Scholar
  13. Pilkis, S. J., El-Maghrabi, M. R., Pilkis, J., Claus, T. H., and Cumming, D. A., 1981, Fructose 2,6-Bisphosphate. A New Activator of Phosphofructokinase, J. Biol. Chem., 256:3171.PubMedGoogle Scholar
  14. Rawitch, A. B., Hudson, E., and Weber, G., 1969, The Rotational Diffusion of Thyroglobulin, J. Biol. Chem., 244:6543.PubMedGoogle Scholar
  15. Reinhart, G. D., 1983, The Determination of Thermodynamic Allosteric Parameters of an Enzyme Undergoing Steady-State Turnover, Arch. Biochem. Biophys., 224:389.PubMedCrossRefGoogle Scholar
  16. Reinhart, G. D., 1985, Influence of pH on the Regulatory Kinetics of Rat Liver Phosphofructokinase: A Thermodynamic Linked-Function Analysis, Biochemistry, 24:7166.PubMedCrossRefGoogle Scholar
  17. Reinhart, G. D., and Hartleip, S. B., 1986, Relationship between Fructose 2,6-Bisphosphate Activation and MgATP Inhibition of Rat Liver Phosphofructokinase at High pH. Kinetic Evidence for Individual Binding Sites Linked by Finite Couplings, Biochemistry, 25:7308.PubMedCrossRefGoogle Scholar
  18. Reinhart, G. D., and Lardy, H. A., 1980a, Rat Liver Phosphofructokinase: Kinetic Activity under Near-Physiological Conditions, Biochemistry, 19:1477.PubMedCrossRefGoogle Scholar
  19. Reinhart, G. D., and Lardy, H. A., 1980b, Rat Liver Phosphofructokinase: Use of Fluorescence Polarization to Study Aggregation at Low Protein Concentration, Biochemistry, 19:1484.PubMedCrossRefGoogle Scholar
  20. Reinhart, G. D., and Lardy, H. A., 1980c, Rat Liver Phosphofructokinase: Kinetic and Physiological Ramifications of the Aggregation Behavior, Biochemistry, 25:1491.CrossRefGoogle Scholar
  21. Sturtevant, J. M., 1977, Heat Capacity and Entropy Changes in Processes Involving Proteins, Proc. Natl. Acad. Sci. U.S.A., 74:2236.PubMedCrossRefGoogle Scholar
  22. Uyeda, K., Furuya, E., and Luby, L. J., 1981a, The Effect of Natural and Synthetic D-Fructose 2,6-Bisphosphate on the Regulatory Kinetic Properties of Liver Phosphofructokinase, J. Biol. Chem., 256:8394.PubMedGoogle Scholar
  23. Uyeda, K., Furuya, E., and Sherry, A. D., 1981b, The Structure of “Activation Factor” for Phosphofructokinase, J. Biol. Chem., 256:8679.PubMedGoogle Scholar
  24. Van Schaftingen, E., Jett, M. F., Hue, L., and Hers, H. G., 1981, Control of Liver Phosphofructokinase by Fructose 2,6-Bisphosphate and Other Effectors, Proc. Natl. Acad. Sci. U.S.A., 78:3483.PubMedCrossRefGoogle Scholar
  25. Weber, G., 1952a, Polarization of the Fluorescence of Macromolecules. I. Theory and Experimental Method, Biochem. J., 51:145.PubMedGoogle Scholar
  26. Weber, G., 1952b, Polarization of the Fluorescence of Macromolecules. II. Fluorescent Conjugates of Ovalbumin and Bovine Serum Albumin, Biochem. J., 51:155.PubMedGoogle Scholar
  27. Weber, G., 1971, The Binding of Small Ligands by Proteins, in: “Horm. Steroids: Proc. 3rd Internatl. Congr.”, Excerpta Medica, Amsterdam, p. 58.Google Scholar
  28. Weber, G., 1972, Ligand Binding and Internal Equilibria in Proteins, Biochemistry, 11:864.PubMedCrossRefGoogle Scholar
  29. Weber, G., 1975, Energetics of Ligand Binding to Proteins, Adv. Prot. Chem. 29:1.CrossRefGoogle Scholar
  30. Wyman, J., 1948, Herne Proteins, Adv. Prot. Chem., 4:407.CrossRefGoogle Scholar
  31. Wyman, J., 1964, Linked Functions and Reciprocal Effects in Hemoglobin: A Second Look, Adv. Prot. Chem., 19:223.CrossRefGoogle Scholar
  32. Wyman, J., 1967, Allosteric Linkage, J. Amer. Chem. Soc., 89:2202.CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1989

Authors and Affiliations

  • Gregory D. Reinhart
    • 1
  1. 1.Department of ChemistryUniversity of OklahomaNormanUSA

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