title: Dissociation of yeast hexokinase by hydrostatic pressure.
abstract: The pressure-induced dissociation of the isozymes P1 and P2 of hexokinase was investigated by studies of the spectral shift of the intrinsic protein fluorescence and by the fluorescence polarization of dansyl conjugates. The free energy of association of the monomers at atmospheric pressure, Katm, was -14.2 kcal mol-1 at 20 degrees C and -11.4 kcal mol-1 at 0 degrees C. The positive enthalpy indicates that the association of the monomers is entropy-driven, overcoming the negative enthalpy of hydration of the subunit interfaces. At 0 degrees C and 1 bar, glucose stabilizes the association by -1.1 kcal mol-1 and the binding of both adenosine 5'-(beta, gamma-methylenetriphosphate) (AMPPCP) and glucose by an even larger amount, -1.34 kcal mol-1. Paradoxically, adenosine 5'-triphosphate (ATP), or AMPPCP, in the absence of glucose destabilizes the association by +0.34 kcal mol-1, while adenosine 5'-diphosphate (ADP) stabilizes it by -0.6 kcal mol-1. Comparison of dV0, the apparent standard volume of association, at different pHs and temperatures indicates that its value (115-160 mL mol-1) is strongly dependent upon the ionization of a group at the subunit interface with a pK near neutrality. Under dissociating pressures, trypsin action results in permanent dissociation of the dimer, confirming earlier observations of Colowick by less direct methods. The P1 and P2 enzymes differ in Katm and dV0 and markedly so in the effects of salt upon the stability of the dimer.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Biochemistry, School of Chemical Sciences, University of Illinois, Urbana 61801.
authors.forename: K
authors.initials: K
authors.lastname: Ruan
authors.forename: G
authors.initials: G
authors.lastname: Weber
mesh_terms: D006593:Hexokinase / Q000378:metabolism*; D006874:Hydrostatic Pressure; D007527:Isoenzymes / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013050:Spectrometry, Fluorescence
chemical_list: D007527:Isoenzymes; D046911:Macromolecular Substances; D006593:Hexokinase