The hexameric ATPase p97 has been implicated in diverse cellular processes through interactions with many different adaptor proteins at its N-terminal domain. Among these, the Ufd1-Npl4 heterodimer is a major adaptor, and the p97-Ufd1-Npl4 complex plays an essential role in endoplasmic reticulum-associated degradation (ERAD), acting as a segregase that translocates the ubiquitinated client protein from the ER membrane into the cytosol for proteasomal degradation. The Nc lobe of the p97 N domain binds specifically to the SHP box motif of Ufd1. The binding interface between the p97 N domain and the Ufd1 SHP motif has been structurally determined. By using mutations of the binding interface residues in the two proteins, we confirmed their functional implication in ERAD. We further addressed a potential regulation of p97-Ufd1 interaction by Ufd1 phosphorylation based on the conserved phosphorylated residues in the Ufd1SHP. Our results suggest that S229 phosphorylation of Ufd1 may negatively regulate the p97N-Ufd1SHP interaction. We also found that a phosphomimetic mutant (S229D) was less effective in mediating degradation of a misfolded protein tyrosinase-C89R, resulting in accumulation of the ERAD substrate. This implies that S229 phosphorylation of Ufd1 may modulate ERAD by impeding p97-Ufd1 interaction. We are currently working on the identification of a relevant protein kinase responsible for the S229 phosphorylation.