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Sci Adv. 2017 Apr 14;3(4):e1700325. doi: 10.1126/sciadv.1700325. eCollection 2017 Apr.

Mechanism of Vps4 hexamer function revealed by cryo-EM.

Author information

1
Life Sciences Institute, University of Michigan, Ann Arbor, MI 48109, USA.
2
Department of Computational Medicine and Bioinformatics, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
3
Department of Biophysics, University of Michigan, Ann Arbor, MI 48109, USA.
4
Department of Chemistry, University of Michigan, Ann Arbor, MI 48109, USA.
5
Department of Biological Chemistry, University of Michigan Medical School, Ann Arbor, MI 48109, USA.

Abstract

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 in multiple 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 substrate-binding loop from the top to the bottom of the central pore, with an associated translation of 33 Å. 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.

KEYWORDS:

AAA+ ATPases; ESCRT III; ESCRT pathway; cryoEM; vps4 structure

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