Mechanistic insights to DNA ligases through biochemical and structural studies

DNA ligases are ubiquitous enzymes dedicated to DNA break repair. My work has focused on modes of DNA damage recognition by exemplary members of the two DNA ligase superfamilies: ATP-dependent ligases and NAD+-dependent ligases. Chlorella virus DNA ligase is the smallest eukaryal ATP-dependent ligase known (298-aa), displays pluripotent biological activity, and has an intrinsic nick-sensing function. Escherichia coli LigA, is the founder of the NAD+-dependent ligase clade, which are essential in all bacteria. We’ve captured crystal structures of these two enzymes at various stages of the nick sealing reaction pathway, which provide deep insights to phosphoryl transfer chemistry and nick recognition, and clues to the design of LigA inhibitors as antibacterial agents. A sub family of ATP-dependent ligases are dedicated to repair of DNA double strand breaks by nonhomologous end joining (NHEJ). Ligase D (Lig D) is the central agent of NHEJ in bacteria. LigD is a multifunctional protein composed of three catalytic modules: ligase, polymerase, and 3’-phosphoesterase (PE). The PE module performs manganese-dependent 3’ end-healing reactions at DSB ends. We’ve determined the atomic structure of Pseudomonas LigD PE at 1.9 Å ,which reveals a novel enzyme fold and a distinctive active site. The PE structure illuminates the catalytic mechanism and the basis for manganese specificity. LigD PE exemplifies a new superfamily of DNA repair enzymes present in all phylogenetic domains.