Structure of the Rpn11-Rpn8 dimer reveals mechanisms of substrate deubiquitination during proteasomal degradation.
Polyubiquitin chains target protein substrates to the 26S proteasome, where they are removed by the deubiquitinase Rpn11 to allow efficient substrate degradation. Despite Rpn11's essential function during substrate processing, its detailed structural and biochemical characterization has been hindered by difficulties in purifying the isolated enzyme. Here we report the 2.0-Å crystal structures of Zn(2+)-free and Zn(2+)-bound Saccharomyces cerevisiae Rpn11 in an MPN-domain heterodimer with Rpn8. The Rpn11-Rpn8 interaction occurs via two distinct interfaces that may be conserved in related MPN-domain complexes. Our structural and mutational studies reveal that Rpn11 lacks a conserved surface to bind the ubiquitin Ile44 patch, does not interact with the moiety on the proximal side of the scissile isopeptide bond and exhibits no linkage specificity for ubiquitin cleavage. These findings explain how Rpn11 functions as a promiscuous deubiquitinase for cotranslocational substrate deubiquitination during proteasomal degradation.
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
Evan J
EJ
Worden
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
Chris
C
Padovani
1] Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA. [2] California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, California, USA.
Andreas
A
Martin
D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D010450:Endopeptidases / Q000737:chemistry* / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D046988:Proteasome Endopeptidase Complex / Q000737:chemistry* / Q000235:genetics; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D025801:Ubiquitin / Q000737:chemistry; D015032:Zinc / Q000737:chemistry
C115451:RPN11 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; D010450:Endopeptidases; D046988:Proteasome Endopeptidase Complex; C573321:Rpn8 protein, S cerevisiae; D015032:Zinc