Symmetry elements in DNA structure important for recognition/methylation by DNA [amino]-methyltransferases

Abstract

The phage T4Dam and EcoDam DNA-[adenine-N6] methyltransferases (MTases) methylate GATC palindromic sequences, while the BamHI DNA-[cytosine-N4] MTase methylates the GGATCC palindrome (which contains GATC) at the internal cytosine residue. We compared the ability of these enzymes to interact productively with defective duplexes in which individual elements were deleted on one chain. A sharp decrease in kcat was observed for all three enzymes if a particular element of structural symmetry was disrupted. For the BamHI MTase, integrity of the ATCC was critical, while an intact GAT sequence was necessary for the activity of T4Dam, and an intact GA was necessary for EcoDam. Theoretical alignment of the region of best contacts between the protein and DNA showed that in the case of a palindromic interaction site, a zone covering the 5'-symmetric residues is located in the major groove versus a zone of contact covering the 3'-symmetric residues in the minor groove. Our data fit a simple rule of thumb that the most important contacts are aligned around the methylation target base: if the target base is in the 5' half of the palindrome, the interaction between the enzyme and the DNA occurs mainly in the major groove; if it is in the 3' half, the interaction occurs mainly in the minor groove.

Figure 1

Effect of deleting an individual nucleotide (A, C and E) or phosphate (B, D and F) in the (bottom strand) recognition site on the kinetics of methyl transfer by the BamHI (A and B) EcoDam (C and D) and T4Dam (E and F) MTases. The data for BamHI were taken from Table 2; the data for EcoDam and T4Dam were taken from previous studies (13,14) in our laboratory.

Figure 2

Theoretical (rectangles) and experimental (ovals) left-to-right alignment of the region of best contacts between the enzyme and the substrate DNA shown in ladder format.