A V-type c subunit in an archaeal ATP synthase: Probing the Na+ binding site in the rotor

A₁A_O ATP synthases are the energy producers in the third domain of life, the archaea. These enzymes are unique and share properties with both, the bacterial F₁F_O ATP synthases and the eukaryal V₁V_O ATPases. Most of the archaea have an F-type like proteolipid (c subunit) in their rotor rings, consisting of one hairpin with one ion binding site. In contrast to that some other archaea, e. g. Pyrococcus furiosus, have an unususal proteolipid (1). The c subunit from this hyperthermophilic archaeon was isolated by chloroform/methanol extraction, purified and the molecular mass was determined with MALDI-TOF-MS. The proteolipid of P. furiosus is a 16 kDa protein, arosen by gene duplication, with two hairpins and two putative ion binding sites. Labeling of the ion binding sites with the ATP synthase inhibitor N,N -Dicyclohexylcarbodiimide (DCCD) clearly showed, that only one ion binding site is functional and that the labeling process is pH-dependent. The effect of Na⁺ on the labeling is object of the current research. Based on these results and the recent finding, that the A₁A_O ATP synthase of P. furiosus has a rotor with 10 c subunits (2), a homology model of the c ring is present, giving first insights into the structure of an archaeal rotor ring with its ion binding site.