DNA methyltransferase Dnmt1 is the key enzyme in functional organization of the human genome. For almost 30 years, Dnmt1 has been unsuccessfully targeted in different drug-design efforts. Most of those failures can be attributed to inadequate understanding of diverse mechanisms that regulate the enzyme activity. Dnmt1 is a large enzyme with multiple flexible domains and phosphorylation sites. Dnmt1 can functionally interact with about 40 different molecules involved in DNA repair, chromatin organization, or RNA-directed-DNA methylation. The interacting molecules include proteins, poly(ADP-ribose), and methylated and unmethylated single-stranded and double-stranded non-coding RNA molecules.We are developing inhibitors and activators of Dnmt1 using a combination of enzyme assays, computational structural analysis, and numerical evaluation of enzyme activity. Knowledge of dynamic reversible processes in the active site of Dnmt1 were used to create novel mechanism-based inhibitors that do not cause DNA damage. Molecular docking and QM/MM MD simulations were used to optimize inhibitors' geometry, flexibility, interaction with the solvent in the active site, and finally formation of a reversible covalent adduct with the enzyme. Different strategies for optimization of the PK/PD properties have been suggested. A dynamic active site loop has been identified as a part of the lock mechanism that controls allosteric regulation and processivity. Enzymatic assays and numerical methods have been developed to study processivity on the DNA substrate and the affinity for binding at the allosteric site. The presented approach allows screening for specific allosteric activators and inhibitors.

DNA methyltransferase Dnmt1: Regulation and novel drug-design strategies

248th National Meeting (2014)

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