Structure of a Ubiquitin E1-E2 Complex: Insights to E1-E2 Thioester Transfer

Ubiquitin (Ub) conjugation is initiated by an E1 enzyme that catalyzes carboxy-terminal Ub adenylation, thioester bond formation to a catalytic cysteine in the E1 Cys domain, and thioester transfer to a catalytic cysteine in E2 conjugating enzymes. How the E1 and E2 active sites come together during...

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Bibliographic Details
Published in:Molecular cell 2013-03, Vol.49 (5), p.884-896
Main Authors: Olsen, Shaun K., Lima, Christopher D.
Format: Article
Language:English
Subjects:
active sites
Adenosine Triphosphate - chemistry
Adenosine Triphosphate - metabolism
Amino Acid Sequence
Catalytic Domain
crystal structure
Crystallography, X-Ray
cysteine
Cysteine - chemistry
Cysteine - metabolism
disulfide bonds
enzymes
Models, Molecular
Protein Structure, Tertiary
Schizosaccharomyces - enzymology
Schizosaccharomyces - metabolism
Schizosaccharomyces pombe Proteins - chemistry
Schizosaccharomyces pombe Proteins - metabolism
ubiquitin
Ubiquitin - chemistry
Ubiquitin - metabolism
Ubiquitin-Conjugating Enzymes - chemistry
Ubiquitin-Conjugating Enzymes - metabolism
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Summary:Ubiquitin (Ub) conjugation is initiated by an E1 enzyme that catalyzes carboxy-terminal Ub adenylation, thioester bond formation to a catalytic cysteine in the E1 Cys domain, and thioester transfer to a catalytic cysteine in E2 conjugating enzymes. How the E1 and E2 active sites come together during thioester transfer and how Ub E1 interacts with diverse Ub E2s remains unclear. Here we present a crystal structure of a Ub E1-E2(Ubc4)/Ub/ATP⋅Mg complex that was stabilized by induction of a disulfide bond between the E1 and E2 active sites. The structure reveals combinatorial recognition of the E2 by the E1 ubiquitin-fold domain (UFD) and Cys domain and mutational analysis, coupled with thioester transfer assays with E1, Ubc4, and other Ub E2s, show that both interfaces are important for thioester transfer. Comparison to a Ub E1/Ub/ATP⋅Mg structure reveals conformational changes in the E1 that bring the E1 and E2 active sites together. ► Crystal structure of ubiquitin E1 in complex with ubiquitin and the E2, Ubc4 ► E1 conformational changes accompany Ub E1-E2 thioester transfer ► Combinatorial recognition of E2 by the E1 Cys and Ubiquitin-fold domains
ISSN:1097-2765
1097-4164
DOI:10.1016/j.molcel.2013.01.013