Ankyrin is a target of spectrin's E2/E3 ubiquitin-conjugating/ligating activity.

T. L. Chang, F. F. Cubillos, D. G. Kakhniashvili, Steven Goodman

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Ubiquitin is a small protein of 8.6 kDa molecular weight. When polyubiquitin is attached to target proteins, they are tagged for destruction by cytoplasmic organelles called proteasomes. We now know that ubiquitination of target proteins also regulates functions as diverse as the sorting of proteins to different intracellular destinations, cell signaling, cell division, gene transcription, and protein-protein interactions. The ubiquitination of target proteins requires a cascade of enzymes: E1 ubiquitin activating enzyme, E2 ubiquitin conjugating enzyme and E3 ubiquitin ligating enzyme. Recently we have demonstrated that the red blood cell (RBC) membrane skeletal protein, spectrin, has E2/E3 enzymatic activities in its alpha-subunit, that can transfer ubiquitin to itself. We have now created a cell free assay using biotinylated ubiquitin that allows detection of target proteins by streptavidin peroxidase. This approach coupled with immunoprecipitation, purification and micro liquid chromatography coupled to tandem mass spectrometry has identified ankyrin as a target of spectrin's E2/E3 activity. Western blotting, with ubiquitin antibody, of purified ankyrin and its well characterized functional domains, has demonstrated that both the spectrin and band 3 binding domains are ubiquitinated in vivo.

Original languageEnglish (US)
Pages (from-to)59-66
Number of pages8
JournalCellular and molecular biology (Noisy-le-Grand, France)
Volume50
Issue number1
StatePublished - Jan 1 2004

Fingerprint

Ankyrins
Spectrin
Ubiquitin
Proteins
Ubiquitination
Ubiquitin-Activating Enzymes
Polyubiquitin
Ubiquitin-Conjugating Enzymes
Streptavidin
Protein Transport
Cell signaling
Proteasome Endopeptidase Complex
Enzymes
Tandem Mass Spectrometry
Immunoprecipitation
Liquid Chromatography
Organelles
Cell Division
Peroxidase
Liquid chromatography

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Ankyrin is a target of spectrin's E2/E3 ubiquitin-conjugating/ligating activity. / Chang, T. L.; Cubillos, F. F.; Kakhniashvili, D. G.; Goodman, Steven.

In: Cellular and molecular biology (Noisy-le-Grand, France), Vol. 50, No. 1, 01.01.2004, p. 59-66.

Research output: Contribution to journalArticle

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