RESEARCH TOPICS

The research in our group focuses on realistic, first-principles simulations of key biological processes of relevance to DNA replication and repair and centers on the following three areas.

Molecular description of archaeal and eukaryotic clamp loading. Proliferating cell nuclear antigen (PCNA) is a toroidal-shaped protein (sliding clamp) that can encircle and slide along DNA. By tethering polymerases to DNA, it ensures processive replication. PCNA plays a pivotal role in DNA replication, modification and repair by coordinating the ordered exchange of repair enzymes and intermediates at the replication fork. To perform its vital functions, the clamp has to be opened and resealed at DNA primer-template junctions by the action of the clamp loader ATPase RFC. Understanding the mechanism of this process constitutes a significant piece in the puzzle of processive DNA replication. Our research aims to provide a unified molecular-level description of prokaryotic, archaeal and eukaryotic clamp loading that would account for all experimental observations available to date.

Conformational switching and exchange of repair enzymes bound to PCNA. In addition to its function as a polymerase accessory factor, PCNA is a recognized master coordinator of cellular responses to DNA damage. Thus, it is of great interest to examine the reversible associations of DNA repair proteins with PCNA and the conformational switching leading to exchange of repair intermediates. Specifically, we will focus on the interactions of flap endonuclease-1 (FEN-1) and DNA ligase 1 (Lig-1) with PCNA and DNA in the context of a proposed handoff mechanism for DNA repair enzymes at the PCNA locus. Groundbreaking structural biology work has opened up the possibility of investigating these large supramolecular assemblies in atomic detail. A modular multi-scale approach will be adopted wherein the individual components of the assembly are first studied by atomistic simulations, followed by construction of lower resolution (coarse grain) models for the complexes. The models could be instrumental in contributing testable hypotheses and directing future experimental efforts in this exciting research area.

Enzyme participants in base excision repair. As a third research thrust, we explore the catalytic repertoire of the enzyme participants in the base excision repair (BER) pathway – a major contributor to genomic integrity in cells. Rationalizing the origins of the remarkable catalytic power of enzymes has been a long-standing challenge for both experimental and theoretical chemical biology. Advances on the theoretical front have been based on the advent of quantum mechanics molecular mechanics (QM/MM) methods. More recently these have been coupled to ab initio molecular dynamics (AIMD) techniques, which have made possible the introduction of dynamical, many-body and polarization effects into simulation models of unprecedented size and sophistication. I will apply state-of-the-art AIMD methodologies to elucidate the origin of the catalytic power of enzymes involved in DNA replication and repair. Detailed mechanistic analysis would reveal the origins of damage recognition and selectivity in these systems. In practical terms, this research has the potential to aid in the development of enzyme inhibitors and thus has direct relevance to drug design. Ultimately, the scope of this research will be expanded to encompass all steps in base excision repair – from the damage specific action of DNA glycosylases, to the final repair step carried out by DNA ligase.

In summary, the research outlined herein is aimed at addressing critical problems in the biomedical arena and unified by the common theme of how cells accomplish faithful duplication of their genetic material. Specifically, the research plan has direct bearing on understanding the molecular basis of genetic integrity and the loss of this integrity in cancer and in degenerative diseases.