American Chemical Society, Journal of the American Chemical Society, 28(133), p. 10708-10711, 2011
DOI: 10.1021/ja202799r
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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 delta-thiol-L-lysine (7) and delta-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.