Assembly, Analysis and Architecture of Atypical Ubiquitin Chains

Nat Struct Mol Biol. 2013 May;20(5):555-65. doi: 10.1038/nsmb.2547. Epub 2013 Apr 7.


Ubiquitin (Ub) chains regulate many cellular processes, but several chain types including Lys6 linkages have remained unstudied. Here we analyze the bacterial effector E3 ligase NleL (non-Lee-encoded effector ligase) from enterohemorrhagic Escherichia coli (EHEC) O157:H7, which assembles Lys6- and Lys48-linked Ub polymers. Using linkage-specific human deubiquitinases (DUBs) we show that NleL generates heterotypic Ub chains, and branched chains are efficiently hydrolyzed by DUBs. USP family DUBs cleave Lys6-linked polymers exclusively from the distal end, whereas a DUB with preference for Lys6 can cleave Lys6-linked polymers at any position in the chain. We used NleL to generate large quantities of Lys6-linked polyUb. Crystallographic and NMR spectroscopy analyses revealed that an asymmetric interface between Ile44 and Ile36 hydrophobic patches of neighboring Ub moieties is propagated in longer Lys6-linked Ub chains. Interactions via the Ile36 patch can displace Leu8 from the Ile44 patch, leading to marked structural perturbations of Ub.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Crystallography, X-Ray
  • Endopeptidases / metabolism
  • Escherichia coli O157 / chemistry*
  • Escherichia coli O157 / metabolism*
  • Escherichia coli Proteins / chemistry*
  • Escherichia coli Proteins / metabolism*
  • Humans
  • Magnetic Resonance Spectroscopy
  • Models, Molecular
  • Protein Conformation
  • Ubiquitin-Protein Ligases / chemistry*
  • Ubiquitin-Protein Ligases / metabolism*


  • Escherichia coli Proteins
  • NleL protein, E coli
  • Ubiquitin-Protein Ligases
  • Endopeptidases

Associated data

  • PDB/3ZLZ