title: ATP binding to p97/VCP D1 domain regulates selective recruitment of adaptors to its proximal N-domain.
abstract: p97/Valosin-containing protein (VCP) is a member of the AAA-ATPase family involved in many cellular processes including cell division, intracellular trafficking and extraction of misfolded proteins in endoplasmic reticulum-associated degradation (ERAD). It is a homohexamer with each subunit containing two tandem D1 and D2 ATPase domains and N- and C-terminal regions that function as adaptor protein binding domains. p97/VCP is directed to its many different functional pathways by associating with various adaptor proteins. The regulation of the recruitment of the adaptor proteins remains unclear. Two adaptor proteins, Ufd1/Npl4 and p47, which bind exclusively to the p97/VCP N-domain and direct p97/VCP to either ERAD-related processes or homotypic fusion of Golgi fragments, were studied here. Surface plasmon resonance biosensor-based assays allowed the study of binding kinetics in real time. In competition experiments, it was observed that in the presence of ATP, Ufd1/Npl4 was able to compete more effectively with p47 for binding to p97/VCP. By using non-hydrolysable ATP analogues and the hexameric truncated p97/N-D1 fragment, it was shown that binding rather than hydrolysis of ATP to the proximal D1 domain strengthened the Ufd1/Npl4 association with the N-domain, thus regulating the recruitment of either Ufd1/Npl4 or p47. This novel role of ATP and an assigned function to the D1 AAA-ATPase domain link the multiple functions of p97/VCP to the metabolic status of the cell.
authors.affiliation: School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
authors.forename: Wei Sheng
authors.initials: WS
authors.lastname: Chia
authors.forename: Diana Xueqi
authors.initials: DX
authors.lastname: Chia
authors.forename: Feng
authors.initials: F
authors.lastname: Rao
authors.forename: Shoshana
authors.initials: S
authors.lastname: Bar Nun
authors.forename: Susana
authors.initials: S
authors.lastname: Geifman Shochat
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001667:Binding, Competitive; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D005470:Fluorometry; D006868:Hydrolysis; D007700:Kinetics; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D020349:Surface Plasmon Resonance
chemical_list: D009687:Nuclear Proteins; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; C404942:p97 ATPase