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Proc Natl Acad Sci U S A. 2014 Nov 25;111(47):16736-41. doi: 10.1073/pnas.1412152111. Epub 2014 Nov 10.

Peptide and small molecule inhibitors of HECT-type ubiquitin ligases.

Author information

1
Medical Research Council Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge CB2 0QH, United Kingdom.
2
Medical Research Council Laboratory of Molecular Biology, Cambridge Biomedical Campus, Cambridge CB2 0QH, United Kingdom hp@mrc-lmb.cam.ac.uk.

Abstract

The human genome encodes several hundred E3 ubiquitin ligases containing RING domains, and around 28 containing HECT domains. These enzymes catalyze the transfer of ubiquitin from E2 enzyme thioesters to a huge range of substrates and play crucial roles in many cellular functions. This makes them attractive potential therapeutic targets. However, they have proven difficult to inhibit: very few good inhibitors exist for RING domain ligases, and none have been described for HECT ligases. Here we show that bicyclic peptides isolated by phage display [Heinis C, Rutherford T, Freund S, Winter G (2009) Nat Chem Biol. 5(7):502-507] can target the E2 binding sites on the HECT domains of Smurf2, Nedd4, Mule/Huwe1, and WWP1, and thus act as specific inhibitors of these enzymes in vitro. By screening for displacement of one of these peptides from Smurf2, we were able to identify a small molecule, heclin (HECT ligase inhibitor), which inhibits several HECT ligases in tissue culture cells. In vitro, heclin does not block E2 binding but causes a conformational change that results in oxidation of the active site Cys. This demonstrates that HECT domains are potentially druggable and provides molecules that may be of experimental use. Heclin kills HEK293 cells growing in culture, consistent with an essential role for HECT ligase activity in mammalian cells.

KEYWORDS:

HECT ligase; inhibitor; phage display; ubiquitin

PMID:
25385595
PMCID:
PMC4250122
DOI:
10.1073/pnas.1412152111
[Indexed for MEDLINE]
Free PMC Article

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