Structural constraints on autoprocessing of the human nucleoporin Nup98

Nucleoporin Nup98, a 98‐kDa protein component of the nuclear pore complex, plays an important role in both protein and RNA transport. During its maturation process, Nup98 undergoes post‐translational autoproteolysis, which is critical for targeting to the NPC. Here we present high‐resolution crystal structures of the C‐terminal autoproteolytic domains of Nup98 (2.3 Å for the wild type and 1.9 Å for the S864A precursor), and propose a detailed autoproteolysis mechanism through an N–O acyl shift. Structural constraints are found at the autocleavage site, and could thus provide a driving force for autocleavage at the scissile peptide bond. Such structural constraints appear to be generated, at least in part, by anchoring a conserved phenylalanine side chain into a highly conserved hydrophobic pocket at the catalytic site. Our high‐resolution crystal structures also reveal that three highly conserved residues, Tyr866, Gly867, and Leu868, provide most of the interactions between the autoproteolytic domain and the C‐terminal tail. These results suggest that Nup98 may represent a new subtype of protein that utilizes autoprocessing to control biogenesis pathways and intracellular translocation.