Domain alternation and active site remodeling are conserved structural features of ubiquitin E1

Zongyang Lv,Lingmin Yuan,James H Atkison,Grace Aldana-Masangkay,Yuan Chen,Shaun K Olsen

doi:10.1074/jbc.m117.787622

Zongyang Lv, Lingmin Yuan + Show 4 more

Open Access

https://doi.org/10.1074/jbc.m117.787622

Copy DOI

Abstract

E1 enzymes for ubiquitin (Ub) and Ub-like modifiers (Ubls) harbor two catalytic activities that are required for Ub/Ubl activation: adenylation and thioester bond formation. Structural studies of the E1 for the Ubl small ubiquitin-like modifier (SUMO) revealed a single active site that is transformed by a conformational switch that toggles its competency for catalysis of these two distinct chemical reactions. Although the mechanisms of adenylation and thioester bond formation revealed by SUMO E1 structures are thought to be conserved in Ub E1, there is currently a lack of structural data supporting this hypothesis. Here, we present a structure of Schizosaccharomyces pombe Uba1 in which the second catalytic cysteine half-domain (SCCH domain) harboring the catalytic cysteine has undergone a 106° rotation that results in a completely different network of intramolecular interactions between the SCCH and adenylation domains and translocation of the catalytic cysteine 12 Å closer to the Ub C terminus compared with previous Uba1 structures. SCCH domain alternation is accompanied by conformational changes within the Uba1 adenylation domains that effectively disassemble the adenylation active site. Importantly, the structural and biochemical data suggest that domain alternation and remodeling of the adenylation active site are interconnected and are intrinsic structural features of Uba1 and that the overall structural basis for adenylation and thioester bond formation exhibited by SUMO E1 is indeed conserved in Ub E1. Finally, the mechanistic insights provided by the novel conformational snapshot of Uba1 presented in this study may guide efforts to develop small molecule inhibitors of this critically important enzyme that is an active target for anticancer therapeutics.

Highlights

E1 enzymes for ubiquitin (Ub) and Ub-like modifiers (Ubls) harbor two catalytic activities that are required for Ub/Ubl activation: adenylation and thioester bond formation
The structural and biochemical data suggest that domain alternation and remodeling of the adenylation active site are interconnected and are intrinsic structural features of Uba1 and that the overall structural basis for adenylation and thioester bond formation exhibited by small ubiquitin-like modifier (SUMO) E1 is conserved in Ub E1
In all structures of canonical E1s determined with their respective cognate Ub/Ubl noncovalently bound to the adenylation domains, the SCCH domain adopts an “open” conformation in which the catalytic cysteine that serves as the nucleophile during thioester bond formation is located more than 30 Å away from C terminus of the Ub/Ubl (14 –16, 18 –22)

Summary

Introduction

E1 enzymes for ubiquitin (Ub) and Ub-like modifiers (Ubls) harbor two catalytic activities that are required for Ub/Ubl activation: adenylation and thioester bond formation. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. Recent structural and biochemical studies revealed that SUMO E1 has a single active site that is reconfigured for catalysis of adenylauitin fold domain; FCCH domain, first catalytic cysteine half-domain; SCCH domain, second catalytic cysteine half-domain; SUMO, small ubiquitin-like modifier; Uba1SCCH_ALT, Uba1 “closed” conformation; Uba1SCCH_OPEN, Uba open conformation (Protein Data Bank code 4II3); SUMO E1SCCH_CLOSED, SUMO E1 closed conformation (Protein Data Bank code 3KYD); SUMO E1SCCH_OPEN, SUMO E1 open conformation (Protein Data Bank code 3KYC); Ub(a), ubiquitin adenylate (i.e. ubiquitin adenylated at its C-terminal glycine residue)

Objectives

Results

Conclusion