Footprint analysis of M.Sssl and M.Hhal methyltransferases reveals extensive interactions with the substrate DNA backbone.
The interactions of the CpG methyltransferases M.Sssl and M.Hhal (GCGC) with substrate DNA were investigated using three different footprinting techniques. The two structurally related enzymes displayed similar specific and non-specific contacts with DNA while bound to their target sequences. DNase I footprinting implicated a region of 18 to 21 base-pairs with which these enzymes interact. Dimethylsulfate protection experiments mostly revealed specific base interactions; each enzyme was shown to interact predominantly with bases at its recognition site in the major groove. However, hydroxyl radical footprints demonstrated extensive interactions with the sugar-phosphate backbone on both strands of the DNA substrate. Both enzymes protected a 16 nucleotide region, in a staggered fashion, covering 9 to 10 nucleotides on each strand. The protected regions, extending for almost a full turn of DNA on each strand, were offset by 6 to 7 nucleotides in the 5' direction, placing both regions on the same face of the double helix, bracketing the major groove. The results suggest that these methyltransferases straddle the major groove from the backbone, but protrude into the groove only to specifically interact with their recognition sites. The sequence-independent interactions observed on the sugar-phosphate backbone may explain the ability of the enzymes to recognize a small sequence, as well as their processive mode of action.
Renbaum
P
P
Department of Cellular Biochemistry, Hebrew University-Hadassah Medical School, Jerusalem, Israel.
Razin
A
A
D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D015257:DNA-Cytosine Methylases / Q000378:metabolism*; D003850:Deoxyribonuclease I; D015226:Dinucleoside Phosphates; D005609:Free Radicals; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000138:chemical synthesis / Q000737:chemistry* / Q000378:metabolism*; D013379:Substrate Specificity
D004247:DNA; D015257:DNA-Cytosine Methylases; D003850:Deoxyribonuclease I; D015226:Dinucleoside Phosphates; D005609:Free Radicals; D009838:Oligodeoxyribonucleotides; C015772:CpG dinucleotide; C037664:DNA modification methylase HhaI; C081573:DNA modification methylase SssI