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Updated: Jul 20, 2026

Ubiquitin Chain Analysis by Parallel Reaction Monitoring
Published on: June 17, 2020
Mms2-Ubc13 covalently bound to ubiquitin reveals the structural basis of linkage-specific polyubiquitin chain
Michael J Eddins1, Candice M Carlile, Kamila M Gomez
1Department of Biophysics and Biophysical Chemistry and the Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
Abstract:
Lys63-linked polyubiquitin chains participate in nonproteolytic signaling pathways, including regulation of DNA damage tolerance and NF-kappaB activation. E2 enzymes bound to ubiquitin E2 variants (UEV) are vital in these pathways, synthesizing Lys63-linked polyubiquitin chains, but how these complexes achieve specificity for a particular lysine linkage has been unclear. We have determined the crystal structure of an Mms2-Ubc13-ubiquitin (UEV-E2-Ub) covalent intermediate with donor ubiquitin linked to the active site residue of Ubc13. In the structure, the unexpected binding of a donor ubiquitin of one Mms2-Ubc13-Ub complex to the acceptor-binding site of Mms2-Ubc13 in an adjacent complex allows us to visualize at atomic resolution the molecular determinants of acceptor-ubiquitin binding. The structure reveals the key role of Mms2 in allowing selective insertion of Lys63 into the Ubc13 active site and suggests a molecular model for polyubiquitin chain elongation.
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