Structure of PvuII DNA (cytosine N4) methyltransferase, an example of domain permutation and protein fold assignment

We have determined the structure of PvuII methyltransferase (M.PvuII) complexed with S-adenosyl-L-methionine (AdoMet) by multiwavelength anomalous diffraction, using a crystal of the selenomethionine-substituted protein. M.PvuII catalyzes transfer of the methyl group from AdoMet to the exocyclic amino (N4) nitrogen of the central cytosine in ifs recognition sequence 5'-CAGCTG-3'. The protein is dominated by an open alpha/beta-sheet structure with a prominent V-shaped cleft: AdoMet and catalytic amino acids are Located at the bottom of this cleft. The size and the basic nature of the cleft are consistent with duplex DNA binding. The target (methylatable) cytosine, if flipped out of the double helical DNA as seen for DNA methyltransferases that generate 5-methylcytosine, would fit into the concave active site next to the AdoMet. This M.PvuII alpha/beta-sheet structure Is very similar to those of M.HhaI (a cytosine C5 methyltransferase) and M.TaqI (an adenine N6 methyltransferase), consistent with a model predicting that DNA methyltransferases share a common structural fold while having the major functional regions permuted into three distinct Linear orders, The main feature of the common fold is a seven-stranded beta-sheet (6 down arrow 7 up arrow 5 down arrow 4 down arrow 1 down arrow 2 down arrow 3 down arrow) formed by five parallel beta-strands and an antiparallel beta-hairpin, The beta-sheet is flanked by six parallel alpha-helices, three on each side. The AdoMet binding site is located at the C-terminal ends of strands beta 1 and beta 2 and the active site is at the C-terminal ends of strands beta 4 and beta 5 and the N-terminal end of strand beta 7., The AdoMet-protein interactions are almost identical among M.PvuII, M.HhaI and M.TaqI, as well as in an RNA methyltransferase and at least one small molecule methyltransferase. The structural similarity among the active sites of M.PvuII, M.TaqI and M.HhaI reveals that catalytic amino acids essential for cytosine N4 and adenine N6 methylation coincide spatially with those for cytosine C5 methylation, suggesting a mechanism for amino methylation.