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J Am Chem Soc. 2011 Jul 20;133(28):10708-11. doi: 10.1021/ja202799r. Epub 2011 Jun 28.

Traceless and site-specific ubiquitination of recombinant proteins.

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  • 1Medical Research Council Laboratory of Molecular Biology, Hills Road, Cambridge CB2 0QH, UK.

Abstract

Protein ubiquitination is a post-translational modification that regulates almost all aspects of eukaryotic biology. Here we discover the first routes for the efficient site-specific incorporation of δ-thiol-L-lysine (7) and δ-hydroxy-L-lysine (8) into recombinant proteins, via evolution of a pyrrolysyl-tRNA synthetase/tRNA(CUA) pair. We combine the genetically directed incorporation of 7 with native chemical ligation and desulfurization to yield an entirely native isopeptide bond between substrate proteins and ubiquitin. We exemplify this approach by demonstrating the synthesis of a ubiquitin dimer and the first synthesis of ubiquitinated SUMO.

PMID:
21710965
PMCID:
PMC3135006
DOI:
10.1021/ja202799r
[PubMed - indexed for MEDLINE]
Free PMC Article
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