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

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.

SUBMITTER: Zhao B

PROVIDER: S-EPMC4104569 | biostudies-literature | 2012 Oct

REPOSITORIES: biostudies-literature

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

Zhao Bo B Bhuripanyo Karan K Zhang Keya K Kiyokawa Hiroaki H Schindelin Hermann H Yin Jun J

Chemistry & biology 20121001 10

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, ...[more]

PMID: 23102221

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