A Mutant of Uracil DNA Glycosylase That Distinguishes between Cytosine and 5-Methylcytosine

作者：Kimber Scott T; Brown Tom; Fox Keith R^*

来源：PLos One, 2014, 9(4): e95394.

DOI：10.1371/journal.pone.0095394

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.

• 出版日期2014-4-16