Determination of Methylation Site of DNA-Methyltransferase NlaX by a Hybrid Method

Abstract

Using a new method based on a combination of bisulfite reaction, the repair enzyme uracil-DNA glycosylase, and synthetic oligodeoxyribonucleotides, the methylation site of DNA-methyltransferase NlaX (M·NlaX) from Neisseria lactamica was established to be the inner cytosine in the double-stranded pentanucleotide recognition sequence 5′-CCNGG-3′ (where N = any nucleoside). 5-Methylcytosine (m5C) type modification by M·NlaX was confirmed by the use of oligonucleotide substrates that contain 5-fluoro-2′-deoxycytidine.