Aroop Sircar - Mechanism of polyubiquitin chain recognition by the human ubiquitin conjugating enzyme Ube2g2

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  Publication Details (including relevant citation   information):

  William E. Bocik, Aroop Sircar, Jeffrey J. Gray & Joel R.   Tolman

  The Journal of Biological Chemistry, 286(5), 3981-3991, February   4, 2011


  Ube2g2 is a human ubiquitin conjugating (E2) enzyme involved   in  the endoplasmic reticulum-associated degradation   pathway, which is  responsible for the identification and   degradation of unfolded and  misfolded proteins in the   endoplasmic reticulum compartment. The  Ube2g2-specific role   is the assembly of Lys-48-linked polyubiquitin  chains,   which constitutes a signal for proteasomal degradation when    attached to a substrate protein. NMR chemical shift perturbation   and  paramagnetic relaxation enhancement approaches were   employed to  characterize the binding interaction between   Ube2g2 and ubiquitin,  Lys-48-linked diubiquitin, and   Lys-63-linked diubiquitin. Results  demonstrate that   ubiquitin binds to Ube2g2 with an affinity of 90 ?M in  two   different orientations that are rotated by 180° in models   generated  by the RosettaDock modeling suite. The binding of   Ube2g2 to Lys-48- and  Lys-63-linked diubiquitin is   primarily driven by interactions with  individual ubiquitin   subunits, with a clear preference for the subunit    containing the free Lys-48 or Lys-63 side chain (i.e. the   distal  subunit). This preference is particularly striking   in the case of  Lys-48-linked diubiquitin, which exhibits an   ?3-fold difference in  affinities between the two ubiquitin   subunits. This difference can be  attributed to the partial   steric occlusion of the subunit whose Lys-48  side chain is   involved in the isopeptide linkage. As such, these results    suggest that Lys-48-linked polyubiquitin chains may be designed   to bind  certain proteins like Ube2g2 such that the terminal   ubiquitin subunit  carrying the reactive Lys-48 side chain   can be positioned properly for  chain elongation regardless   of chain length.

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