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. 2013 Mar;9(3):154-6.
doi: 10.1038/nchembio.1159. Epub 2013 Jan 6.

A conserved asparagine has a structural role in ubiquitin-conjugating enzymes

Christopher E Berndsen  1 , Reuven Wiener, Ian W Yu, Alison E Ringel, Cynthia Wolberger
Affiliations

A conserved asparagine has a structural role in ubiquitin-conjugating enzymes

Christopher E Berndsen et al. Nat Chem Biol. 2013 Mar.

Abstract

It is widely accepted that ubiquitin-conjugating enzymes contain an active site asparagine that serves as an oxyanion hole, thereby stabilizing a negatively charged transition state intermediate and promoting ubiquitin transfer. Using structural and biochemical approaches to study the role of the conserved asparagine to ubiquitin conjugation by Ubc13-Mms2, we conclude that the importance of this residue stems primarily from its structural role in stabilizing an active site loop.

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Conflict of interest statement

Competing Financial Interests Statement

The Authors declare no competing financial interests.

Figures

Figure 1
Figure 1. Polyubiquitin chain formation by Ubc13/Mms2 in the presence and absence of Rad5
(a) Substrate partitioning experiments showing active fraction of Ubc13~Ub thioester as a function of acceptor ubiquitin concentration. Plot of percentage diubiquitin formed versus concentration of ubiquitin (Ub)Δ 75,Δ76 for Ubc13N79Q-Mms2 (filled circles) or Ubc13N79Q-Mms2 with the Rad5 RING domain (open circles). Points represent the average of 3 to 4 separate measurements with the standard deviation shown by the error bars. (b) Single discharge assays of diubiquitin formation by Ubc13-Mms2 containing wild-type Ubc13 or mutants with substitutions at N79, performed in the presence and absence of the Rad5 RING fragment. Full gels in Supplementary Figure 4.
Figure 2
Figure 2. Crystal structure of Ubc13N79A
(a) Alignment of the structure of Ubc13N79A (salmon) with wild-type Ubc13 from 2GMI (yellow). Residue names are colored to match the coloring of the N79A or wild-type structures. (b) Electron density 2F0-Fc map contoured at 1.0σ showing the density for the active site loop in Ubc13N79A. The backbone Cα trace of wild-type Ubc13 from 2GMI is shown in green. (c) Single discharge assay for the Ubc13 active site loop mutants using ubiquitinΔ75,Δ76 (Ub) as the ubiquitin acceptor.
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References

    1. Pickart CM, Eddins MJ. Ubiquitin: structures, functions, mechanisms. Biochim Biophys Acta. 2004;1695:55–72. - PubMed
    1. Kerscher O, Felberbaum R, Hochstrasser M. Modification of proteins by ubiquitin and ubiquitin-like proteins. Annu Rev Cell Dev Biol. 2006;22:159–80. - PubMed
    1. Wu PY, et al. A conserved catalytic residue in the ubiquitin-conjugating enzyme family. EMBO J. 2003;22:5241–50. - PMC - PubMed
    1. Yunus AA, Lima CD. Lysine activation and functional analysis of E2-mediated conjugation in the SUMO pathway. Nat Struct Mol Biol. 2006;13:491–9. - PubMed
    1. Wenzel DM, Stoll KE, Klevit RE. E2s: structurally economical and functionally replete. Biochem J. 2010;433:31–42. - PMC - PubMed

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