2.3 Å resolution cryo-EM structure of human p97 and mechanism of allosteric inhibition

Abstract

p97 is a hexameric AAA ATPase that is an attractive target for cancer drug development. Here, we report cryo-EM structures for ADP-bound, full-length, hexameric wild-type p97 in the presence and absence of an allosteric inhibitor at resolutions of 2.3 Å and 2.4 Å, respectively. We also report cryo-EM structures at ~ 3.3 Å, 3.2 Å and 3.3 Å resolutions respectively, for three distinct, co-existing functional states of p97 with occupancies of 0, 1 or 2 molecules of ATPγS per protomer. A large corkscrew-like change in molecular architecture coupled with upward displacement of the N-domain is observed only when ATPγS is bound to both D1 and D2 domains. These cryo-EM structures establish the sequence of nucleotide-driven structural changes in p97 at atomic resolution. They also enable elucidation of the binding mode of an allosteric small molecule inhibitor to p97 and illustrate how inhibitor binding at the interface between D1 and D2 domains prevents propagation of the conformational changes necessary for p97 function