Analysis of protein ubiquitination.

Jeffrey D. Laney, Mark Hochstrasser

Research output: Contribution to journalArticle

1 Scopus citations

Abstract

Attachment of ubiquitin (Ub) to a protein requires a complex of enzymes that recognize the substrate and promote Ub transfer. Sequence motifs present in these enzymes may indicate that other uncharacterized proteins containing these motifs have a biochemical function of Ub-protein ligation, and several in vitro methods are described in this unit for determining if a protein has Ub-transferring activity. They include psmunoblotting of psmunoprecipitated proteins, affinity purification using His-tagged ubiquitin, assaying for auto-ubiquitination of E3, and assaying ubiquitination of a model substrate protein. These methods are suitable for a variety of eukaryotic cells, but techniques are specifically described for use with yeast and mammalian cells.

Original languageEnglish (US)
Pages (from-to)Unit 14.5
JournalCurrent protocols in protein science / editorial board, John E. Coligan ... [et al.]
VolumeChapter 14
DOIs
StatePublished - Nov 2002

ASJC Scopus subject areas

  • Structural Biology
  • Biochemistry

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