Abstract
E1 enzymes activate ubiquitin (Ub) and ubiquitin-like modifiers (Ubls) in the first step of Ub/Ubl conjugation cascades and represent potential targets for therapeutic intervention in cancer and other life-threatening diseases. Here, we report the crystal structure of the E1 enzyme for the Ubl SUMO in complex with a recently discovered and highly specific covalent allosteric inhibitor (COH000). The structure reveals that COH000 targets a cryptic pocket distinct from the active site that is completely buried in all previous SUMO E1 structures and that COH000 binding to SUMO E1 is accompanied by a network of structural changes that altogether lock the enzyme in a previously unobserved inactive conformation. These structural changes include disassembly of the active site and a 180° rotation of the catalytic cysteine-containing SCCH domain, relative to conformational snapshots of SUMO E1 poised to catalyze adenylation. Altogether, our study provides a molecular basis for the inhibitory mechanism of COH000 and its SUMO E1 specificity, and also establishes a framework for potential development of molecules targeting E1 enzymes for other Ubls at a cryptic allosteric site.
Highlights
E1 enzymes activate ubiquitin (Ub) and ubiquitin-like modifiers (Ubls) in the first step of Ub/ Ubl conjugation cascades and represent potential targets for therapeutic intervention in cancer and other life-threatening diseases
The E1 enzymes for all Ubls share conserved structural features including the adenylation domain (AAD), inactive adenylation domain (IAD), and catalytic cysteine residues, and all activate their cognate Ubl via a two-step catalytic mechanism involving sequential adenylation and thioester bond formation 29-38Recent studies have revealed that both the adenylation and thioester bond formation activities of SUMO E1 are catalyzed at a single location on the enzyme that is reconfigured for catalysis of these distinct chemical reactions via a network of complementary conformational changes[27]
When in complex with COH000, SUMO E1 residues essential for adenylation are either disordered or displaced from the active site and the second catalytic cysteine half-domain (SCCH) domain undergoes a 180° rotation that is stabilized by a new network of contacts with the adenylation domains that lock the E1 in an inactive state
Summary
E1 enzymes activate ubiquitin (Ub) and ubiquitin-like modifiers (Ubls) in the first step of Ub/ Ubl conjugation cascades and represent potential targets for therapeutic intervention in cancer and other life-threatening diseases. The structure reveals that COH000 targets a cryptic pocket distinct from the active site that is completely buried in all previous SUMO E1 structures and that COH000 binding to SUMO E1 is accompanied by a network of structural changes that altogether lock the enzyme in a previously unobserved inactive conformation These structural changes include disassembly of the active site and a 180° rotation of the catalytic cysteine-containing SCCH domain, relative to conformational snapshots of SUMO E1 poised to catalyze adenylation. The E1 enzymes for all Ubls share conserved structural features including the AAD, IAD, and catalytic cysteine residues, and all activate their cognate Ubl via a two-step catalytic mechanism involving sequential adenylation and thioester bond formation 29-38Recent studies have revealed that both the adenylation and thioester bond formation activities of SUMO E1 are catalyzed at a single location on the enzyme that is reconfigured for catalysis of these distinct chemical reactions via a network of complementary conformational changes[27]. E1s for other Ubls harbor pockets analogous to the COH000 binding site of SUMO E1, raising the possibility that these sites can be exploited for the development of inhibitors targeting other Ubl pathways
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