title: Human DNMT1 transition state structure.
abstract: Human DNA methyltransferase 1 (DNMT1) maintains the epigenetic state of DNA by replicating CpG methylation signatures from parent to daughter strands, producing heritable methylation patterns through cell divisions. The proposed catalytic mechanism of DNMT1 involves nucleophilic attack of Cys(1226) to cytosine (Cyt) C6, methyl transfer from S-adenosyl-l-methionine (SAM) to Cyt C5, and proton abstraction from C5 to form methylated CpG in DNA. Here, we report the subangstrom geometric and electrostatic structure of the major transition state (TS) of the reaction catalyzed by human DNMT1. Experimental kinetic isotope effects were used to guide quantum mechanical calculations to solve the TS structure. Methyl transfer occurs after Cys(1226) attack to Cyt C6, and the methyl transfer step is chemically rate-limiting for DNMT1. Electrostatic potential maps were compared for the TS and ground states, providing the electronic basis for interactions between the protein and reactants at the TS. Understanding the TS of DNMT1 demonstrates the possibility of using similar analysis to gain subangstrom geometric insight into the complex reactions of epigenetic modifications.
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461.
authors.forename: Quan
authors.initials: Q
authors.lastname: Du
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Department of Biochemistry, Albert Einstein College of Medicine, Bronx, NY 10461 vern.schramm@einstein.yu.edu.
authors.forename: Vern L
authors.initials: VL
authors.lastname: Schramm
mesh_terms: D001667:Binding, Competitive; D002247:Carbon Isotopes / Q000737:chemistry; D002250:Carbon Radioisotopes / Q000737:chemistry; D002384:Catalysis; D003545:Cysteine / Q000737:chemistry; D003596:Cytosine / Q000737:chemistry / Q000378:metabolism; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000737:chemistry* / Q000378:metabolism; D019175:DNA Methylation; D003903:Deuterium / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012435:S-Adenosylhomocysteine / Q000378:metabolism; D014316:Tritium / Q000737:chemistry
chemical_list: D002247:Carbon Isotopes; D002250:Carbon Radioisotopes; D011993:Recombinant Fusion Proteins; D014316:Tritium; D003596:Cytosine; D012435:S-Adenosylhomocysteine; D003903:Deuterium; D000076123:DNA (Cytosine-5-)-Methyltransferase 1; D004248:DNA (Cytosine-5-)-Methyltransferases; C000619380:DNMT1 protein, human; D003545:Cysteine
keywords: 5-methylcytosine; CpG methylation; DNA methyltransferase; S-adenosyl-l-methionine; transition state