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Chembiochem. 2016 Jun 2;17(11):995-8. doi: 10.1002/cbic.201600042. Epub 2016 Apr 26.

Chemical Protein Ubiquitylation with Preservation of the Native Cysteine Residues.

Author information

1
Department of Chemistry and Biochemistry, University of Delaware, 214A Drake Hall, Newark, DE, 19716, USA.
2
Department of Chemistry and Biochemistry, University of Delaware, 214A Drake Hall, Newark, DE, 19716, USA. zzhuang@udel.edu.

Abstract

We report a cysteine-based ligation strategy for generating a monoubiquitylated protein while preserving the native cysteine residues on the acceptor protein. In monoubiquitylation of proliferating cell nuclear antigen (PCNA) this method circumvents the need to mutate the native cysteine residues on PCNA. The chemically ubiquitylated PCNA contains a noncleavable linkage of the same length as the native isopeptide linkage. It also retains the normal function of the native Ub-PCNA in stimulating the ATPase activity of replication factor C (RFC) and lesion bypass synthesis by Polη. This method may be adapted for chemical ubiquitylation of other proteins and for site-specific modification of a target protein at a specific site through sulfhydryl chemistry.

KEYWORDS:

caged cysteine; chemical ligation; photo-deprotection; protecting groups; protein modifications; ubiquitylation

PMID:
27113245
PMCID:
PMC5298353
DOI:
10.1002/cbic.201600042
[Indexed for MEDLINE]
Free PMC Article

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