Grant: Discovery Grants Program - Individual

Double Strand Breaks (DSBs) represent the most extreme form of damage experienced by the genome, resulting in genetic loss and/or chromosomal re-arrangement if not properly repaired. Exogenous (radiation and various chemicals) and endogenous (DNA replication and repair) sources of damage are capable of generating lethal DSBs. Given the potential loss of genetic information, it is not surprising that tolerable levels of exposure to damage such as ionizing radiation are extremely small for all forms of life. One notable exception is the bacterium Deinococcus radiodurans, which has the remarkable ability to withstand doses of ionizing radiation 1500 times greater than humans can survive and one that would obliterate the E. coli genome 10 times over. Remarkably, despite being shattered into hundreds of small DNA fragments, Deinococcus radiodurans readily reassembles a functional genome. This feat has been attributed to the combination of a number of protective mechanisms including the presence of multiple genome copies, characteristic genome structure, and known or novel DNA repair mechanisms. Although traditional repair mechanisms appear to be active in D. radiodurans, the speed and accuracy of repair displayed are inconsistent with all previously characterized pathways. The goal of our research is to understand how uncharacterized proteins, implicated in DNA repair, function to permit the amazing DNA repair/assembly capabilities of Deinococcus radiodurans. The mechanistic basis of repair will be analyzed for 2 of the 5 known proteins that that are unique to Deinococci and essential for resistance to extreme levels of DNA damage. Proposed studies of DdrB and DdrC include structural and biophysical characterization, as well as functional analysis involving in vitro (DNA interaction, repair complex assembly) and in vivo (localization, survival) studies. Understanding the DNA repair strategies utilized to resist extreme IR damage is not only fascinating from a scientific point of view, but also carries great potential for practical utility. Deinococcus represent a new chassis for industrial biotechnology and are of interest for applications such as bioremediation of toxins from radioactive contaminated environments generated through processing of nuclear fuel. In addition, NASA is exploring these organisms with the idea of eventually using information gained to increase protection form cosmic radiation during prolonged space travel. The greatest impact, however, that understanding (and exploiting) DNA repair/assembly capabilities of D. radiodurans resides in the area of synthetic biology. The next frontier in biological sciences is, without doubt, the ability to engineer chromosomes at the truly synthetic level. A major challenge to such endeavors is the ability to efficiently and accurately assemble large, chromosome sized DNA molecules. Deinococci are well equipped to overcome this limitation.