Amino-terminal dimerization, NRDP1-rhodanese interaction, and inhibited catalytic domain conformation of the ubiquitin-specific protease 8 (USP8)

J Biol Chem. 2006 Dec 8;281(49):38061-70. doi: 10.1074/jbc.M606704200. Epub 2006 Oct 11.

Abstract

Ubiquitin-specific protease 8 (USP8) hydrolyzes mono and polyubiquitylated targets such as epidermal growth factor receptors and is involved in clathrin-mediated internalization. In 1182 residues, USP8 contains multiple domains, including coiled-coil, rhodanese, and catalytic domains. We report the first high-resolution crystal structures of these domains and discuss their implications for USP8 function. The amino-terminal domain is a homodimer with a novel fold. It is composed of two five-helix bundles, where the first helices are swapped, and carboxyl-terminal helices are extended in an antiparallel fashion. The structure of the rhodanese domain, determined in complex with the E3 ligase NRDP1, reveals the canonical rhodanese fold but with a distorted primordial active site. The USP8 recognition domain of NRDP1 has a novel protein fold that interacts with a conserved peptide loop of the rhodanese domain. A consensus sequence of this loop is found in other NRDP1 targets, suggesting a common mode of interaction. The structure of the carboxyl-terminal catalytic domain of USP8 exhibits the conserved tripartite architecture but shows unique traits. Notably, the active site, including the ubiquitin binding pocket, is in a closed conformation, incompatible with substrate binding. The presence of a zinc ribbon subdomain near the ubiquitin binding site further suggests a polyubiquitin-specific binding site and a mechanism for substrate induced conformational changes.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Catalytic Domain
  • Crystallography, X-Ray
  • Dimerization
  • Endopeptidases / chemistry*
  • Endopeptidases / genetics
  • Endopeptidases / metabolism*
  • Endosomal Sorting Complexes Required for Transport
  • Humans
  • In Vitro Techniques
  • Kinetics
  • Models, Molecular
  • Multiprotein Complexes
  • Protein Structure, Quaternary
  • Protein Structure, Tertiary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Static Electricity
  • Thiosulfate Sulfurtransferase / chemistry*
  • Thiosulfate Sulfurtransferase / genetics
  • Thiosulfate Sulfurtransferase / metabolism*
  • Ubiquitin Thiolesterase
  • Ubiquitin-Protein Ligases / chemistry*
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*

Substances

  • Endosomal Sorting Complexes Required for Transport
  • Multiprotein Complexes
  • Recombinant Proteins
  • RNF41 protein, human
  • Ubiquitin-Protein Ligases
  • Thiosulfate Sulfurtransferase
  • Endopeptidases
  • USP8 protein, human
  • Ubiquitin Thiolesterase
  • ubiquitin isopeptidase

Associated data

  • PDB/1A9U
  • PDB/1WFW
  • PDB/1WHB
  • PDB/2FZP
  • PDB/2GFO
  • PDB/2GWF