Mechanism of Vps4 hexamer function revealed by cryo-EM

Vps4 is a member of AAA(+) ATPase (adenosine triphosphatase associated with diverse cellular activities) that operates as an oligomer to disassemble ESCRT-III (endosomal sorting complex required for transport III) filaments, thereby catalyzing the final step inmultiple ESCRT-dependent membrane remodeling events. We used electron cryo-microscopy to visualize oligomers of a hydrolysis-deficient Vps4 (vacuolar protein sorting- associated protein 4) mutant in the presence of adenosine 5'-triphosphate (ATP). We show that Vps4 subunits assemble into an asymmetric hexameric ring following an approximate helical path that sequentially stacks substrate-binding loops along the central pore. The hexamer is observed to adopt an open or closed ring configuration facilitated by major conformational changes in a single subunit. The structural transition of the mobile Vps4 subunit results in the repositioning of its substratebinding loop fromthe top to the bottom of the central pore, with an associated translation of 33 Alpha. These structures, along with mutant-doping experiments and functional assays, provide evidence for a sequential and processive ATP hydrolysis mechanism by which Vps4 hexamers disassemble ESCRT-III filaments.