A mutant of uracil DNA glycosylase that distinguishes between cytosine and 5-methylcytosine.

Journal article

We demonstrate that a mutant of uracil DNA glycosylase (N123D:L191A) distinguishes between cytosine and methylcytosine. Uracil DNA glycosylase (UDG) efficiently removes uracil from DNA in a reaction in which the base is flipped into the enzyme's active site. Uracil is selected over cytosine by a pattern of specific hydrogen bonds, and thymine is excluded by steric clash of its 5-methyl group with Y66. The N123D mutation generates an enzyme that excises cytosine. This N123D:L191A mutant excises C when it is mispaired with A or opposite an abasic site, but not when it is paired with G. In contrast no cleavage is observed with any substrates that contain 5-methylcytosine. This enzyme may offer a new approach for discriminating between cytosine and 5-methylcytosine.

Authors: Kimber, S; Brown, T; Fox, K

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Public Library of Science
• Journal: PloS one
• Volume: 9
• Issue: 4
• Pages: e95394
• Publication date: 2014-01-01
• Acceptance date: 2014-03-25
• DOI: 10.1371/journal.pone.0095394
• EISSN: 1932-6203
• ISSN: 1932-6203
• Language: English
• Keywords: Thymine; Uracil-DNA Glycosidase; Escherichia coli; Bacterial Proteins; Substrate Specificity; Base Sequence; Uracil; Mutation; DNA; Oligonucleotides; Gene Expression; 5-Methylcytosine; Recombinant Proteins; Cytosine; Molecular Sequence Data