DNA methyltransferases: epigenetics, enzyme mechanisms, and the development of novel therapeutics

S-Adenosyl methionine-dependent DNA methyltransferases modify DNA sequence-specifically. Bacterial DNA is modified at both cytosine (N4 or C5) and adenine (N6), while mammalian DNA is modified only at cytosine C5. The bacterial enzymes are involved in restriction/modification, mismatch repair, replication timing, and gene regulation. These enzymes are essential to the virulence of several human pathogens. Numerous high resolution x-ray structures are available for these bacterial enzymes, enabling our efforts to understand the following aspects:

1. Mechanisms of catalysis
2. Importance of conformational mechanisms, including DNA bending, base flipping, intercalation, and protein loop motion towards catalysis and specificity.
3. The importance of correlated protein motion to catalysis, and the use of such information to identify allosteric sites which may form the basis of designing allosteric inhibitors.
4. Antibiotic design based on inhibiting the bacterial DNA MTases.
5. Determining how DNA methylation regulates virulence in the major causative microbe which causes periodontitis, Actinobacillus actinomycetomcomitans.
6. The engineering of new specificities through the combined use of unnatural amino acids, artificial cell sorting, and in vitro evolution.

Mammalian DNA methylation is involved in parent of origin imprinting, x-chromosome inactivation, host defense, and tissue-specific gene regulation. Aberrant DNA methylation of tumor suppressor genes is a primary and early event in human tumorigenesis. A major unresolved question is how do the patterns of DNA methylation, which occur in the CpG dinucleotide, become established? Recent work shows that the mammalian DNA cytosine methyltransferases (Dnmt1, Dnmt2, Dnmt3 family) interact with other cellular proteins, and that de novo DNA methylation is directed in part by non-coding RNA. Our efforts are focused on studying the human enzymes to understand how interactions with other proteins and RNA direct the function and specificity of these enzymes. We have also identified inhibitors of the mammalian enzymes which are cell active.

Nanoparticle-based proteomics, diagnostics, and basic cell biology tools

Modern molecular biology techniques allow the creation of templates and materials of very exacting reactivity, size, structure, and periodicity. These materials can be usefully coupled to inorganic compounds to create bio-materials that are active and well-controlled on a molecular scale. Currently, our investigations are most concerned with utilizing the programmability and specificity of DNA, coupled with the ability to stoichiometrically decorate homogeneous metallic nanoparticles, towards the creation of novel bio-sensor and nano-electronics components with tightly controlled electrical and optical properties.