Vfa1 Binds to the N-terminal Microtubule Interacting and Trafficking (MIT) Domain of Vps4 and Stimulates Its ATPase Activity [Protein Structure and Folding]

February 24th, 2014 by Vild, C. J., Xu, Z.

The endosomal sorting complexes required for transport (ESCRTs) are responsible for multivesicular body (MVB) biogenesis, membrane abscission during cytokinesis, and retroviral budding. They function as transiently-assembled molecular complexes on the membrane and their disassembly requires the action of the AAA-ATPase Vps4. Vps4 is regulated by a multitude of ESCRT and ESCRT-related proteins. Binding of these proteins to Vps4 is often mediated via the microtubule interacting and trafficking (MIT) domain of Vps4. Recently, a new Vps4-binding protein Vfa1 was identified in a yeast genetic screen, where overexpression of Vfa1 caused defects in vacuolar morphology. However, the function of Vfa1 and its role in vacuolar biology were largely unknown. Here, we provide a first detailed biochemical and biophysical study of Vps4-Vfa1 interaction. The MIT domain of Vps4 binds to the C-terminal seventeen residues of Vfa1. This interaction is of high affinity and greatly stimulates the ATPase activity of Vps4. The crystal structure of the Vps4-Vfa1 complex shows that Vfa1 adopts a canonical MIT-interacting Motif 2 (MIM2) structure that has been previously observed in other Vps4-ESCRT interactions. These findings suggest that Vfa1 is a novel positive regulator of Vps4 function.
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