A systems-wide screen identifies substrates of the SCFβTrCP ubiquitin ligase

Low Teck Yew, Mao Peng, Roberto Magliozzi, Shabaz Mohammed, Daniele Guardavaccaro, Albert J R Heck

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

Cellular proteins are degraded by the ubiquitin-proteasome system (UPS) in a precise and timely fashion. Such precision is conferred by the high substrate specificity of ubiquitin ligases. Identification of substrates of ubiquitin ligases is crucial not only to unravel the molecular mechanisms by which the UPS controls protein degradation but also for drug discovery purposes because many established UPS substrates are implicated in disease. We developed a combined bioinformatics and affinity purification-mass spectrometry (AP-MS) workflow for the system-wide identification of substrates of SCFβTrCP, a member of the SCF family of ubiquitin ligases. These ubiquitin ligases are characterized by a multisubunit architecture typically consisting of the invariable subunits Rbx1, Cul1, and Skp1, and one of 69 F-box proteins. The F-box protein of thismember of the family is βTrCP. SCFβTrCPbinds, through theWD40 repeats of βTrCP, to the DpSGXX(X)pS diphosphorylated motif in its substrates. We recovered 27 previously reported SCFβTrCP substrates, of which 22 were verified by two independent statistical protocols, thereby confirming the reliability of this approach. In addition to known substrates, we identified 221 proteins that contained the DpSGXX(X)pS motif and also interacted specifically with the WD40 repeats of βTrCP. Thus, with SCFβTrCP, as the example, we showed that integration of structural information, AP-MS, and degron motif mining constitutes an effective method to screen for substrates of ubiquitin ligases.

Original languageEnglish
Article numberrs8
JournalScience Signaling
Volume7
Issue number356
DOIs
Publication statusPublished - 1 Jan 2014
Externally publishedYes

Fingerprint

Ligases
Ubiquitin
Substrates
Proteasome Endopeptidase Complex
F-Box Proteins
Mass Spectrometry
SKP Cullin F-Box Protein Ligases
Purification
Mass spectrometry
Workflow
Proteins
Drug Discovery
Substrate Specificity
Computational Biology
Bioinformatics
Proteolysis
Control systems
Degradation

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology
  • Medicine(all)

Cite this

Teck Yew, L., Peng, M., Magliozzi, R., Mohammed, S., Guardavaccaro, D., & Heck, A. J. R. (2014). A systems-wide screen identifies substrates of the SCFβTrCP ubiquitin ligase. Science Signaling, 7(356), [rs8]. https://doi.org/10.1126/scisignal.2005882

A systems-wide screen identifies substrates of the SCFβTrCP ubiquitin ligase. / Teck Yew, Low; Peng, Mao; Magliozzi, Roberto; Mohammed, Shabaz; Guardavaccaro, Daniele; Heck, Albert J R.

In: Science Signaling, Vol. 7, No. 356, rs8, 01.01.2014.

Research output: Contribution to journalArticle

Teck Yew, L, Peng, M, Magliozzi, R, Mohammed, S, Guardavaccaro, D & Heck, AJR 2014, 'A systems-wide screen identifies substrates of the SCFβTrCP ubiquitin ligase', Science Signaling, vol. 7, no. 356, rs8. https://doi.org/10.1126/scisignal.2005882
Teck Yew, Low ; Peng, Mao ; Magliozzi, Roberto ; Mohammed, Shabaz ; Guardavaccaro, Daniele ; Heck, Albert J R. / A systems-wide screen identifies substrates of the SCFβTrCP ubiquitin ligase. In: Science Signaling. 2014 ; Vol. 7, No. 356.
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