title: N6-Adenosine DNA Methyltransferase from H. pylori 98-10 Strain in Complex with DNA and AdoMet: Structural Insights from in Silico Studies.
abstract: Helicobacter pylori is a primitive Gram-negative bacterium that resides in the acidic environment of the human gastrointestinal tract, and some strains of this bacterium cause gastric ulcers and cancer. DNA methyltransferases (MTases) are promising drug targets for the treatment of cancer and other diseases that are also caused by epigenetic alternations of the genome. The N6-adenine-specific DNA MTase from H. pylori (M. Hpy N6mA) catalyzes the transfer of a methyl group from the cofactor S-adenosyl-l-methionine (AdoMet) to the flipped adenine of the substrate DNA. In this work, we report the sequence analyses, three-dimensional structure modeling, and molecular dynamics simulations of M. Hpy N6mA, when complexed with AdoMet as well as DNA. We analyzed the protein-DNA interactions prominently established by the flipped cytosine and the interactions between protein cofactors in the active site. The comparable orientation of AdoMet in both systems confirms that AdoMet is in a catalytically competent orientation in the bimolecular system that is retained upon DNA binding in the termolecular system of M. Hpy N6mA. In both systems, AdoMet is stabilized in the binding pocket by hydrogen bonding (Thr84, Glu99, Asp122, and Phe123) as well as van der Waals (Ile100, Phe160, Arg104, and Cys76) interactions. We propose that the contacts made by flipped adenine DA6 with Asn138 (N6 and N1 atom of DA6) and Pro139 (N6) and π-stacking interactions with Phe141 and Phe219 play an important role in the methylation mechanism at the N6 position in our N6mA model. Specific recognition of DNA is mediated by residues 143-155, 183-189, 212-220, 280-293, and 308-325. These findings are further supported by alanine scanning mutagenesis studies. The conserved residues in distantly related sequences of the small domain are important in DNA binding. Results reported here elucidate the sequence, structure, and binding features necessary for the recognition between cofactor AdoMet and substrate DNA by the vital epigenetic enzyme, M. Hpy N6mA.
authors.affiliation: School of Chemistry, University of Hyderabad , Hyderabad 500046, India.
authors.forename: Swati
authors.initials: S
authors.lastname: Singh
authors.affiliation: School of Chemistry, University of Hyderabad , Hyderabad 500046, India.
authors.forename: Lalitha
authors.identifier: 0000-0002-1579-4611
authors.initials: L
authors.lastname: Guruprasad
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D020134:Catalytic Domain; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D016480:Helicobacter pylori; D006860:Hydrogen Bonding; D056004:Molecular Dynamics Simulation; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D012436:S-Adenosylmethionine / Q000737:chemistry / Q000378:metabolism*; D016415:Sequence Alignment; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D012436:S-Adenosylmethionine; D004247:DNA; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)