Functional dissection of a HECT ubiquitin E3 ligase.

Ubiquitination is one of the most prevalent protein post-translational modifications in eukaryotes, and its malfunction is associated with a variety of human diseases. Despite the significance of this process, the molecular mechanisms that govern the regulation of ubiquitination remain largely unknown. Here we used a combination of yeast proteome chip ...
assays, genetic screening, and in vitro/in vivo biochemical analyses to identify and characterize eight novel in vivo substrates of the ubiquitinating enzyme Rsp5, a homolog of the human ubiquitin-ligating enzyme Nedd4, in yeast. Our analysis of the effects of a deubiquitinating enzyme, Ubp2, demonstrated that an accumulation of Lys-63-linked polyubiquitin chains results in processed forms of two substrates, Sla1 and Ygr068c. Finally we showed that the localization of another newly identified substrate, Rnr2, is Rsp5-dependent. We believe that our approach constitutes a paradigm for the functional dissection of an enzyme with pleiotropic effects.
Mesh Terms:
Endopeptidases, Endosomal Sorting Complexes Required for Transport, Humans, Lysine, Polyubiquitin, Protein Processing, Post-Translational, Proteome, Reproducibility of Results, Saccharomyces cerevisiae, Saccharomyces cerevisiae Proteins, Substrate Specificity, Ubiquitin-Protein Ligase Complexes
Mol. Cell Proteomics
Date: Jan. 01, 2008
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