An NMR-based model of the ubiquitin-bound human ubiquitin conjugation complex Mms2.Ubc13. The structural basis for lysine 63 chain catalysis.
A heterodimer composed of the catalytically active ubiquitin-conjugating enzyme hUbc13 and its catalytically inactive paralogue, hMms2, forms the catalytic core for the synthesis of an alternative type of multiubiquitin chain where ubiquitin molecules are tandemly linked to one another through a Lys-63 isopeptide bond. This type of linkage, as opposed to the more typical Lys-48-linked chains, serves as a non-proteolytic marker of protein targets involved in error-free post-replicative DNA repair and NF-kappa B signal transduction. Using a two-dimensional (1)H-(15)N NMR approach, we have mapped: 1) the interaction between the subunits of the human Ubc13.Mms2 heterodimer and 2) the interactions between each of the subunits or heterodimer with a non-covalently bound acceptor ubiquitin or a thiolester-linked donor ubiquitin. Using these NMR-derived constraints and an unbiased docking approach, we have assembled the four components of this catalytic complex into a three-dimensional model that agrees well with its catalytic function.
McKenna
Sean
S
Department of Biochemistry, University of Alberta, Edmonton, Alberta T6G 2H7, Canada.
Moraes
Trevor
T
Pastushok
Landon
L
Ptak
Christopher
C
Xiao
Wei
W
Spyracopoulos
Leo
L
Ellison
Michael J
MJ
D000595:Amino Acid Sequence; D001665:Binding Sites; D002384:Catalysis; D019281:Dimerization; D006801:Humans; D008025:Ligases / Q000737:chemistry* / Q000378:metabolism*; D008239:Lysine; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D011487:Protein Conformation; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D025801:Ubiquitin / Q000737:chemistry* / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes
D025801:Ubiquitin; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases; D008239:Lysine