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Functional dissection of a HECT ubiquitin E3 ligase.

ABSTRACT: 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.

SUBMITTER: Lu JY

PROVIDER: S-EPMC2861892 | biostudies-literature | 2008 Jan

REPOSITORIES: biostudies-literature

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Functional dissection of a HECT ubiquitin E3 ligase.

Lu Jin-Ying JY Lin Yu-Yi YY Qian Jiang J Tao Sheng-Ce SC Zhu Jian J Pickart Cecile C Zhu Heng H

Molecular & cellular proteomics : MCP 20071019 1

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 ubiquitina ...[more]

PMID: 17951556

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