Conformational Changes in Chloroplast F₁-ATPase Caused by Thiol-Dependent Activation and MgADP-Dependent Inactivation

Abstract—The state of tyrosine residues of the chloroplast coupling factor CF₁ was studied by spectrophotometric titration. It was shown that some tyrosine residues of CF₁ underwent deprotonation at pH values of the medium much lower than the pK of free tyrosine. The number of such residues depends on both the conformational state of the enzyme and the composition of the medium. They are abundant in CF₁ with the γ-subunit that contains a disulfide bridge. Bridge reduction leads to a decrease of their number and, accordingly, an increase in the number of residues that undergo deprotonation at a pH higher than the tyrosine pK. The minimum number of residues that dissociated within the 6.0–9.0 pH range was observed in the reaction mixture containing Mg²⁺ or MgADP. It is assumed that the changes in pK values for tyrosine residues result from the presence or absence of positively charged amino-acid residues in their neighborhood, which is indicative of alterations in the tertiary structure of the enzyme. Deprotonation of a considerable part of tyrosine residues in the presence of Mg²⁺ or MgADP occurs within an abnormally narrow pH range and demonstrates the cooperative transition to the new conformational state of the enzyme. Comparison of the data obtained with our previous kinetic data indicates that the titration characteristics and the respective structures of CF₁-ATPase observed in the presence of Mg²⁺ or MgADP result from reversible inactivation caused by MgADP binding to one catalytic site and one noncatalytic site.