New publication in "Nucleic Acids Research"

New publication in "Nucleic Acids Research"

DNA-(cytosine-C5)-methyltransferases (MTases) represent a large group of evolutionary related enzymes with specific DNA interaction. We systematically investigated the specificity and flanking sequence preferences of six bacterial enzymes of this class. For each enzyme, several methylation experiments were conducted, in total 196 experiments comprising more than 1,300,000 individual sequence reads. This allowed us to determine the recognition specificity and flanking sequence preferences (and their mutual connection) quantitatively and with high resolution providing interesting and novel insights into the reaction mechanism of DNA methyltransferases. We observed high (>1000-fold) target sequence specificity reflecting strong evolutionary pressure against unspecific DNA methylation. Strong flanking sequence preferences (~100-fold) were observed. Mutation of amino acids involved in DNA contacts led to local changes of specificity and flanking sequence preferences, but also global effects indicating that larger conformational changes occur upon transition state formation. Based on these findings, we conclude that the transition state of the DNA methylation reaction precedes the covalent enzyme-DNA complex conformations with flipped target base that are resolved in structural studies. Moreover, our data suggest that alternative catalytically active conformations exist whose occupancy is modulated by enzyme-DNA contacts. Sequence dependent DNA shape analyses suggest that MTase flanking sequence preferences are caused by flanking sequence dependent modulation of the DNA conformation. Considering DNA methyltransferases an exemplary, well-studied group of evolutionary related enzymes that specifically interact with DNA, many of our results are likely to be transferable to other DNA interacting enzymes and proteins.