Redox regulation of CF1-ATPase involves interplay between the γ-subunit neck region and the turn region of the βDELSEED-loop

The soluble F-1 complex of ATP synthase (F0F1) is capable of ATP hydrolysis, accomplished by the minimum catalytic core subunits alpha(3)beta(3)gamma. A special feature of cyanobacterial F-1 and chloroplast F-1 (CF1) is an amino acid sequence inserted in the gamma-subunit. The insertion is extended slightly into the CF1 enzyme containing two additional cysteines for regulation of ATPase activity via thiol modulation. This molecular switch was transferred to a chimeric F-1 by inserting the cysteine-containing fragment from spinach CF1 into a cyanobacterial gamma-subunit [Y. Kim et al., redox regulation of rotation of the cyanobacterial F-1-ATPase containing thiol regulation switch, J Biol Chem, 286 (2011) 9071-9078]. Under oxidizing conditions, the obtained F-1 tends to lapse into an ADP-inhibited state, a common regulation mechanism to prevent wasteful ATP hydrolysis under unfavorable circumstances. However, the information flow between thiol modulation sites on the gamma-subunit and catalytic sites on the beta-subunits remains unclear. Here, we clarified a possible interplay for the CF1-ATPase redox regulation between structural elements of the beta DELSEED-loop and the gamma-subunit neck region, i.e., the most convex part of the alpha-helical gamma-termini. Critical residues were assigned on the beta-subunit, which received the conformation change signal produced by disulfide/dithiol formation on the gamma-subunit. Mutant response to the ATPase redox regulation ranged from lost to hypersensitive. Furthermore, mutant cross-link experiments and inversion of redox regulation indicated that the gamma-redox state might modulate the subunit interface via reorientation of the beta DELSEED motif region. (C) 2015 Elsevier B.V. All rights reserved.