title: Thermodynamics of nucleotide binding to the chaperonin GroEL studied by isothermal titration calorimetry: evidence for noncooperative nucleotide binding.
abstract: We characterized the thermodynamics of binding reactions of nucleotides ADP and ATPgammaS (a nonhydrolyzable analog of ATP) to GroEL in a temperature range of 5 degrees C to 35 degrees C by isothermal titration calorimetry. Analysis with a noncooperative binding model has shown that the bindings of nucleotides are driven enthalpically with binding constants of 7x103 M-1 and 4x104 M-1 for ADP and ATPgammaS, respectively, at 26 degrees C and that the heat capacity change DeltaCp is about 100 cal/mol.K for both the nucleotides. The stoichiometries of binding were about 8 and 9 molecules for ADP and ATPgammaS, respectively, per GroEL tetradecamer at 5 degrees C, and both increased with temperature to reach about 14 (ADP) and 12 (ATPgammaS) for both nucleotides at 35 degrees C. The absence of initial increase of binding heat as well as Hill coefficient less than 1.2, which were obtained from the fitting to the model curve by assuming positive cooperativity, showed that there was virtually no positive cooperativity in the nucleotide bindings. Incorporating a difference in affinity for the nucleotide (ADP and ATPgammaS) between the two rings of GroEL into the noncooperative binding model improved the goodness of fitting and the difference in the affinity increased with decreasing temperature.
authors.affiliation: Department of Physics, School of Science, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: T P
authors.initials: TP
authors.lastname: Terada
authors.forename: K
authors.initials: K
authors.lastname: Kuwajima
mesh_terms: D000244:Adenosine Diphosphate / Q000737:chemistry; D000494:Allosteric Regulation; D002151:Calorimetry / Q000379:methods; D018834:Chaperonin 60 / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D004926:Escherichia coli; D008962:Models, Theoretical; D009711:Nucleotides / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D013816:Thermodynamics
chemical_list: D018834:Chaperonin 60; D009711:Nucleotides; D000244:Adenosine Diphosphate