Preferential interaction of sentrin with a ubiquitin-conjugating enzyme, Ubc9

Limin Gong, Tetsu Kamitani, Kenichi Fujise, Laura S. Caskey, Edward T H Yeh

Research output: Contribution to journalArticle

150 Citations (Scopus)

Abstract

Sentrin is a ubiquitin-like molecule that has been shown to interact with the death domains of Fas and tumor necrosis factor receptor 1 (TNFR1), PML, Rad51, Rad52, and RanGAP1. We have reported previously that sentrin can be conjugated to other proteins in a manner analogous to protein ubiquitination (Kamitani, T., Nguyen, H. P., and Yeh, E. T. H. (1997) J. Biol. Chem. 272, 14001-14004). Furthermore, the conserved C-terminal Gly-Gly residues are required for sentrinization to occur. To identify enzymes which play a role in sentrinization, the yeast two-hybrid system was used to screen a human placenta cDNA library using sentrin as bait. A strong positive interacting clone was found to contain a cDNA insert encoding the ubiquitin- conjugating enzyme, Ubc9. The interaction between sentrin and Ubc9 required the ubiquitin domain and the C-terminal Gly-Gly residues of sentrin. This interaction appears to be specific because sentrin could only interact weakly with UbcH5B, but could not interact with HHR6B, UbcH6 nor E2-EPF. In vitro translated sentrin could be precipitated by a GST-Ubc9 fusion protein, but not by glutathione S-transferase. A β-mercaptoethanol-sensitive Ubc9- sentrin conjugate could also be identified in the in vitro binding assay. Substitution of the conserved cysteine residue of Ubc9 by serine abolished the formation of the Ubc9-sentrin conjugate. Taken together, Ubc9 is a strong candidate to be the key conjugating enzyme in the sentrinization pathway.

Original languageEnglish (US)
Pages (from-to)28198-28201
Number of pages4
JournalJournal of Biological Chemistry
Volume272
Issue number45
DOIs
StatePublished - Nov 7 1997
Externally publishedYes

Fingerprint

SUMO-1 Protein
Ubiquitin-Conjugating Enzymes
Ubiquitin
Ubiquitin C
Two-Hybrid System Techniques
Proteins
Mercaptoethanol
Tumor Necrosis Factor Receptors
Ubiquitination
Enzymes
Glutathione Transferase
Hybrid systems
Gene Library
Yeast
Serine
Placenta
Cysteine
Assays
Substitution reactions
Fusion reactions

ASJC Scopus subject areas

  • Biochemistry

Cite this

Preferential interaction of sentrin with a ubiquitin-conjugating enzyme, Ubc9. / Gong, Limin; Kamitani, Tetsu; Fujise, Kenichi; Caskey, Laura S.; Yeh, Edward T H.

In: Journal of Biological Chemistry, Vol. 272, No. 45, 07.11.1997, p. 28198-28201.

Research output: Contribution to journalArticle

Gong, Limin ; Kamitani, Tetsu ; Fujise, Kenichi ; Caskey, Laura S. ; Yeh, Edward T H. / Preferential interaction of sentrin with a ubiquitin-conjugating enzyme, Ubc9. In: Journal of Biological Chemistry. 1997 ; Vol. 272, No. 45. pp. 28198-28201.
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