Characterization of a Pleiotropic Protein Promoting DNA Repair

Abstract

<p>DNA double-strand breaks (DSBs) represent the most severe form of chromosomal damage. In higher organisms, one DSB represents a lethal event. Unlike any other organism, bacteria of the genus <em>Deinococcus</em> are able to withstand extremely high levels of DSBs caused by ionizing radiation, UV radiation, prolonged desiccation and chemical agents. The remarkable ability of <em>Deinococcus</em> to survive under conditions of extreme oxidative stress is a result of an efficient DSB repair process. Currently, little is known about how several key proteins function in DNA repair in <em>Deinococcus</em>. The goal of this work was to better understand the mechanism through which <em>Deinococcus</em> is able to recover from DNA damage through structural and biochemical characterization of a unique pleiotropic protein promoting DNA repair. Here, we show preliminary X-ray diffraction data coupled with electron microscopy images which showcase unusual filament formation of this protein. Pleiotropic protein promoting DNA repair has been shown by others to bind double-stranded DNA (dsDNA). We have further characterized the interaction of this protein with DNA and found that it binds longer pieces of dsDNA (> 3500 bp) with a higher affinity than shorter pieces of dsDNA (≤ 1000 bp). This protein was also shown to form bundles of long, fibrous filaments by transmission electron microscopy, while gel filtration studies have shown it exists as a very large multimer in solution. Implications for how these filaments function in DNA repair were explored. <em>In vivo</em> damage recovery through complementation studies were also performed. Determining the structure of pleiotropic protein promoting DNA repair, and characterizing its DNA binding ability and filament formation is essential to clarify its proposed function in DNA repair.</p>