Structural basis of the interaction between the AAA ATPase p97/VCP and its adaptor protein p47

Abstract

The AAA ATPase p97/VCP is involved in many cellular events including ubiquitin-dependent processes and membrane fusion. In the latter, the p97 adaptor protein p47 is of central importance. In order to provide insight into the molecular basis of p97 adaptor binding, we have determined the crystal structure of p97 ND1 domains complexed with p47 C-terminal domain at 2.9 A resolution. The structure reveals that the p47 ubiquitin regulatory X domain (UBX) domain interacts with the p97 N domain via a loop (S3/S4) that is highly conserved in UBX domains, but is absent in ubiquitin, which inserts into a hydrophobic pocket between the two p97 N subdomains. Deletion of this loop and point mutations in the loop significantly reduce p97 binding. This hydrophobic binding site is distinct from the predicted adaptor-binding site for the p97/VCP homologue N-ethylmaleimide sensitive factor (NSF). Together, our data suggest that UBX domains may act as general p97/VCP/CDC48 binding modules and that adaptor binding for NSF and p97 might involve different binding sites. We also propose a classification for ubiquitin-like domains containing or lacking a longer S3/S4 loop.

Structural insights into the p97-Ufd1-Npl4 complex

“…We have tentatively assigned positions of the Ufd1-Npl4 domains in our projection structure based on size of density and labeling studies with nanogold and antibody. In comparison, p47 has three well defined domains (18)(19)(20)33) separated by long linkers and is a trimer in solution (19,31). Despite p47 (Ϸ150 kDa) being larger than the Ufd1-Npl4 complex, it is not readily visualized by EM (data not shown), probably because of the inherent flexibility of the individual domains.…”

Structural Basis for Ovarian Tumor Domain-containing Protein 1 (OTU1) Binding to p97/Valosin-containing Protein (VCP)

“…This may be due to the additional proline in UBXL OTU1 . Conversely, the proline in the S3/S4 loop of UBX p47 was reported to be in trans-configuration (36). The corresponding region in UBD NPL4 is longer by six residues and forms a 3 10 -helix instead of the loop as shown in Fig.…”

Control of p97 function by cofactor binding

“…Molecular insights into p97-UBX domain interactions were obtained by crystal structures of the N domain in complex with the UBX domains of p47 [53] and FAF1 [54][55][56] (Fig. 2B).…”