title: Designed semisynthetic protein inhibitors of Ub/Ubl E1 activating enzymes.
abstract: Semisynthetic, mechanism-based protein inhibitors of ubiquitin (Ub) and ubiquitin-like modifier (Ubl) activating enzymes (E1s) have been developed to target E1-catalyzed adenylation and thioesterification of the Ub/Ubl C-terminus during the processes of protein SUMOylation and ubiquitination. The inhibitors were generated by intein-mediated expressed protein ligation using a truncated Ub/Ubl protein (SUMO residues 1-94; Ub residues 1-71) with a C-terminal thioester and synthetic tripeptides having a C-terminal adenosine analogue and an N-terminal cysteine residue. SUMO-AMSN (4a) and Ub-AMSN (4b) contain a sulfamide group as a nonhydrolyzable mimic of the phosphate group in the cognate Ub/Ubl-AMP adenylate intermediate in the first half-reaction, and these constructs selectively inhibit SUMO E1 and Ub E1, respectively, in a dose-dependent manner. SUMO-AVSN (5a) and Ub-AVSN (5b) contain an electrophilic vinyl sulfonamide designed to trap the incoming E1 cysteine nucleophile (Uba2 Cys173 in SUMO E1; Uba1 Cys593 in Ub E1) in the second half-reaction, and these constructs selectively, covalently, and stably cross-link to SUMO E1 and Ub E1, respectively, in a cysteine nucleophile-dependent manner. These inhibitors are powerful tools to probe outstanding mechanistic questions in E1 function and can also be used to study the biological functions of E1 enzymes.
authors.affiliation: Molecular Pharmacology and Chemistry Program, Memorial Sloan-Kettering Cancer Center, 1275 York Avenue, Box 422, New York, New York 10065, USA.
authors.forename: Xuequan
authors.initials: X
authors.lastname: Lu
authors.forename: Shaun K
authors.initials: SK
authors.lastname: Olsen
authors.forename: Allan D
authors.initials: AD
authors.lastname: Capili
authors.forename: Justin S
authors.initials: JS
authors.lastname: Cisar
authors.forename: Christopher D
authors.initials: CD
authors.lastname: Lima
authors.forename: Derek S
authors.initials: DS
authors.lastname: Tan
mesh_terms: D015195:Drug Design; D004791:Enzyme Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology*; D008958:Models, Molecular; D011487:Protein Conformation; D025842:SUMO-1 Protein / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism*; D044764:Ubiquitin-Activating Enzymes / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism*; D014452:Ubiquitins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D025842:SUMO-1 Protein; D025801:Ubiquitin; D014452:Ubiquitins; D044764:Ubiquitin-Activating Enzymes