Specificity of the BRISC Deubiquitinating Enzyme Is Not Due to Selective Binding to Lys63-linked Polyubiquitin

BRISC (Brcc36-containing isopeptidase complex) is a four-subunit deubiquitinating (DUB) enzyme that has a catalytic subunit, called Brcc36, that is a member of the JAMM/MPN+ family of zinc metalloproteases. A notable feature of BRISC is its high specificity for cleaving Lys63-linked polyubiquitin. H...

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Bibliographic Details
Published in:	The Journal of biological chemistry 2010-04, Vol.285 (14), p.10344-10352
Main Authors:	Cooper, Eric M., Boeke, Jef D., Cohen, Robert E.
Format:	Article
Language:	eng
Subjects:	Binding Sites Carbon-Nitrogen Lyases - metabolism Catalytic Domain Deubiquitinating Enzymes Enzymes/Kinetics Enzymes/Metallo Enzymes/Proteolytic HeLa Cells Humans Lysine - metabolism Membrane Proteins - metabolism Multiprotein Complexes - genetics Multiprotein Complexes - metabolism Polyubiquitin - metabolism Proteases/Metalloprotease Proteases/Ubiquitin Proteases/Ubiquitination Proteasome Endopeptidase Complex - metabolism Protein Binding Protein Synthesis and Degradation Protein/Post-translational Modification Protein/Turnover Recombinant Proteins - genetics Recombinant Proteins - isolation & purification Recombinant Proteins - metabolism Substrate Specificity Trans-Activators - metabolism Ubiquitination Ubiquitins - metabolism
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Summary:	BRISC (Brcc36-containing isopeptidase complex) is a four-subunit deubiquitinating (DUB) enzyme that has a catalytic subunit, called Brcc36, that is a member of the JAMM/MPN+ family of zinc metalloproteases. A notable feature of BRISC is its high specificity for cleaving Lys63-linked polyubiquitin. Here, we show that BRISC selectivity is not due to preferential binding to Lys63-linked polyubiquitin but is instead dictated by how the substrate isopeptide linkage is oriented within the enzyme active site. BRISC possesses a high affinity binding site for the ubiquitin hydrophobic surface patch that accounts for the bulk of the affinity between enzyme and substrate. Although BRISC can interact with either subunit of a diubiquitin conjugate, substrate cleavage occurs only when BRISC is bound to the hydrophobic patch of the distal (i.e. the “S1”) ubiquitin at a ubiquitin-ubiquitin cleavage site. The importance of the Lys63-linked proximal (S1′) ubiquitin was underscored by our finding that BRISC could not cleave the isopeptide bond joining a ubiquitin to a non-ubiquitin substrate. Finally, we also show that Abro1, another BRISC subunit, binds directly to Brcc36 and that the Brcc36-Abro1 heterodimer includes a minimal complex with Lys63-specific DUB activity.
ISSN:	0021-9258 1083-351X