The Biological Role of the ζ Subunit as Unidirectional Inhibitor of the F1FO-ATPase of Paracoccus denitrificans

Abstract

The biological roles of the three natural F₁F_O-ATPase inhibitors, ε, ζ, and IF₁, on cell physiology remain controversial. The ζ subunit is a useful model for deletion studies since it mimics mitochondrial IF₁, but in the F₁F_O-ATPase of Paracoccus denitrificans (PdF₁F_O), it is a monogenic and supernumerary subunit. Here, we constructed a P. denitrificans 1222 derivative (PdΔζ) with a deleted ζ gene to determine its role in cell growth and bioenergetics. The results show that the lack of ζ in vivo strongly restricts respiratory P. denitrificans growth, and this is restored by complementation in trans with an exogenous ζ gene. Removal of ζ increased the coupled PdF₁F_O-ATPase activity without affecting the PdF₁F_O-ATP synthase turnover, and the latter was not affected at all by ζ reconstitution in vitro. Therefore, ζ works as a unidirectional pawl-ratchet inhibitor of the PdF₁F_O-ATPase nanomotor favoring the ATP synthase turnover to improve respiratory cell growth and bioenergetics.

Keywords: ATP synthase; Paracoccus denitrificans; knockout; nanomotor; oxidative phosphorylation; pawl; ratchet; rotation; unidirectional inhibitor; ζ subunit.