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Issue 10, 2010
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N-Methylcysteine-mediated total chemical synthesis of ubiquitin thioester

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Ubiquitin thioester is a key intermediate in the ubiquitylation of proteins and is formed enzymatically through the activation of α-COOH of ubiquitin in an ATP dependent manner using the E1 enzyme. The current methods used for the preparation of ubiquitin thioester rely on either the enzymatic machinery or on expressed protein ligation technology. In this article, we report a new chemical strategy, combining native chemical ligation and N-methylcysteine containing peptides, to chemically prepare ubiquitin thioester for the first time. The N-methylcysteine is utilized as an N→S acyl transfer device, and in its protected form serves as a latent thioester functionality. This enabled us to trigger the formation of ubiquitin thioester subsequent to the assembly of the ubiquitin polypeptide via native chemical ligation. The synthetic ubiquitin thioester showed a similar behavior in peptide ubiquitylation to the one obtained via expression. This approach should allow for higher flexibility in the chemical manipulation of ubiquitin thioester in a wide variety of ubiquitylated peptides and proteins for structural and biochemical analysis and for the synthesis of ubiquitin chains.

Graphical abstract: N-Methylcysteine-mediated total chemical synthesis of ubiquitin thioester

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Publication details

The article was received on 12 Jan 2010, accepted on 19 Feb 2010 and first published on 11 Mar 2010

Article type: Paper
DOI: 10.1039/C000332H
Citation: Org. Biomol. Chem., 2010,8, 2392-2396

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    N-Methylcysteine-mediated total chemical synthesis of ubiquitin thioester

    L. A. Erlich, K. S. A. Kumar, M. Haj-Yahya, P. E. Dawson and A. Brik, Org. Biomol. Chem., 2010, 8, 2392
    DOI: 10.1039/C000332H

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