Structure of yeast RAVE bound to a partial V₁ complex.

Abstract

Vacuolar-type ATPases (V-ATPases) are membrane-embedded proton pumps that acidify intracellular compartments in almost all eukaryotic cells. Homologous with ATP synthases, these multisubunit enzymes consist of a soluble catalytic V₁ subcomplex and a membrane-embedded proton-translocating V_O subcomplex. The V₁ and V_O subcomplexes can undergo reversible dissociation to regulate proton pumping, with reassociation of V₁ and V_O requiring the protein complex known as RAVE (regulator of the ATPase of vacuoles and endosomes). In the yeast <i>Saccharomyces cerevisiae</i>, RAVE consists of subunits Rav1p, Rav2p, and Skp1p. We used electron cryomicroscopy (cryo-EM) to determine a structure of yeast RAVE bound to V₁. In the structure, RAVE is an L-shaped complex with Rav2p pointing toward the membrane and Skp1p distant from both the membrane and V₁. Only Rav1p interacts with V₁, binding to a region of subunit A not found in the corresponding ATP synthase subunit. When bound to RAVE, V₁ is in a rotational state suitable for binding the free V_O complex, but in the structure, it is partially disrupted, missing five of its 16 subunits. Other than these missing subunits and the conformation of the inhibitory subunit H, the V₁ complex with RAVE appears poised for reassembly with V_O.