title: Tyr-341 of the beta subunit is a major Km-determining residue of TF1-ATPase: parallel effect of its mutations on Kd(ATP) of the beta subunit and on Km(ATP) of the alpha 3 beta 3 gamma complex.
abstract: Residue Tyr-341 of the F1-ATPase beta subunit from a thermophilic Bacillus strain, PS3, was mutagenized to leucine, cysteine or alanine. Each of the mutated beta subunits was isolated and its affinity for ATP-Mg was examined by means of difference circular dichroism and differential titration calorimetry. The Kd values for ATP-Mg obtained were: beta Y341 (wild type), 0.015 mM; beta Y341L, 0.7 mM; beta Y341C and beta Y341A, > 3 mM. All the mutant beta subunits could be reconstituted into the alpha 3 beta 3 gamma complex with alpha and gamma subunits. The alpha 3 beta (mutant)3 gamma complexes hydrolyzed ATP with apparent Vmax values larger than that of the alpha 3 beta (WILD)3 gamma complex. The apparent Km values of the alpha 3 beta (mutant)3 gamma complexes increased in parallel with the Kd values for ATP-Mg of the isolated mutant beta subunits. These results indicate that residue beta Y341 is directly involved in the catalytic ATP-Mg binding and is a major Km-determining residue of F1-ATPase.
authors.affiliation: Chemical Engineering Laboratory, Institute of Physical and Chemical Research, Saitama.
authors.forename: M
authors.initials: M
authors.lastname: Odaka
authors.forename: C
authors.initials: C
authors.lastname: Kaibara
authors.forename: T
authors.initials: T
authors.lastname: Amano
authors.forename: T
authors.initials: T
authors.lastname: Matsui
authors.forename: E
authors.initials: E
authors.lastname: Muneyuki
authors.forename: K
authors.initials: K
authors.lastname: Ogasahara
authors.forename: K
authors.initials: K
authors.lastname: Yutani
authors.forename: M
authors.initials: M
authors.lastname: Yoshida
mesh_terms: D000255:Adenosine Triphosphate / Q000494:pharmacology; D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002151:Calorimetry / Q000379:methods; D002384:Catalysis; D002942:Circular Dichroism; D003545:Cysteine / Q000737:chemistry; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry*; D006180:Proton-Translocating ATPases / Q000737:chemistry*; D013816:Thermodynamics; D017186:Titrimetry; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D010446:Peptide Fragments; D014443:Tyrosine; D000255:Adenosine Triphosphate; D006180:Proton-Translocating ATPases; D003545:Cysteine