Orthogonal ubiquitin transfer through engineered E1-E2 cascades for protein ubiquitination

Bo Zhao, Karan Bhuripanyo, Keya Zhang, Hiroaki Kiyokawa, Hermann Schindelin, Jun Yin*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

17 Scopus citations

Abstract

Protein modification by ubiquitin (UB) controls diverse cellular processes. UB is conjugated to cellular proteins by sequential transfer through an E1-E2-E3 enzymatic cascade. The cross-activities of 2 E1s, 50 E2s and thousands of E3s encoded by the human genome make it difficult to identify the substrate proteins of a specific E3 enzyme in the cell. One way to solve this problem is to engineer an orthogonal UB transfer (OUT) cascade in which the engineered UB (xUB) is relayed by engineered E1, E2 and E3 enzymes (xE1, xE2, xE3) to modify the substrate proteins of a specific E3. Here, we use phage display and mutagenesis to construct xUB-xE1 and xE1-xE2 pairs that are orthogonal to the native E1 and E2 enzymes. Our work on engineering the UB transfer cascades will enable us to use OUT to map the signal transduction networks mediated by protein ubiquitination.

Original languageEnglish (US)
Pages (from-to)1265-1277
Number of pages13
JournalChemistry and Biology
Volume19
Issue number10
DOIs
StatePublished - Oct 26 2012

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Pharmacology
  • Drug Discovery
  • Clinical Biochemistry

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