Selectivity of pyruvate kinase for Na+ and K+ in water/dimethylsulfoxide mixtures

Eur J Biochem. 2003 Jun;270(11):2377-85. doi: 10.1046/j.1432-1033.2003.03605.x.


In aqueous media, muscle pyruvate kinase is highly selective for K+ over Na+. We now studied the selectivity of pyruvate kinase in water/dimethylsulfoxide mixtures by measuring the activation and inhibition constants of K+ and Na+, i.e. their binding to the monovalent and divalent cation binding sites of pyruvate kinase, respectively [Melchoir J.B. (1965) Biochemistry 4, 1518-1525]. In 40% dimethylsulfoxide the K0.5 app for K+ and Na+ were 190 and 64-fold lower than in water. Ki app for K+ and Na+ decreased 116 and 135-fold between 20 and 40% dimethylsulfoxide. The ratios of Ki app/K0.5 app for K+ and Na+ were 34-3.5 and 3.3-0.2, respectively. Therefore, dimethylsulfoxide favored the partition of K+ and Na+ into the monovalent and divalent cation binding sites of the enzyme. The kinetics of the enzyme at subsaturating concentrations of activators show that K+ and Mg2+ exhibit high selectivity for their respective cation binding sites, whereas when Na+ substitutes K+, Na+ and Mg2+ bind with high affinity to their incorrect sites. This is evident by the ratio of the affinities of Mg2+ and K+ for the monovalent cation binding site, which is close to 200. For Na+ and Mg2+ this ratio is approximately 20. Therefore, the data suggest that K+ induces conformational changes that prevent the binding of Mg2+ to the monovalent cation binding site. Circular dichroism spectra of the enzyme and the magnitude of the transfer and apparent binding energies of K+ and Na+ indicate that structural arrangements of the enzyme induced by dimethylsulfoxide determine the affinities of pyruvate kinase for K+ and Na+.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Binding Sites
  • Binding, Competitive
  • Cations
  • Circular Dichroism
  • Dimethyl Sulfoxide / chemistry*
  • Dose-Response Relationship, Drug
  • Ions
  • Kinetics
  • Magnesium / chemistry
  • Potassium / chemistry
  • Potassium / metabolism*
  • Protein Conformation
  • Pyruvate Kinase / chemistry*
  • Rabbits
  • Sodium / chemistry
  • Sodium / metabolism*
  • Spectrometry, Fluorescence
  • Swine
  • Thermodynamics
  • Ultraviolet Rays
  • Water / chemistry


  • Cations
  • Ions
  • Water
  • Sodium
  • Pyruvate Kinase
  • Magnesium
  • Potassium
  • Dimethyl Sulfoxide