Bioinformatic Analysis And Anti-oxidization Function Of The RecD Protein From Deinococcus Radiodurans

The extremely radiation resistant vegetative bacterium Deinococcus radiodurans was isolated in 1956 from gamma-irradiated meat. Since then, it has been the subject of research aimed at understanding the genetic mechanisms underlying its resistance phenotype. In addition to being able to survive high doses of acute gamma radiation, the organism can grow under chronic gamma radiation and recover from a range of other DNA damaging conditions including exposure to desiccation, ultraviolet, hydrogen peroxide and chemical genotoxic agents. This remarkable range of resistance has been attributed to efficient DNA repair processes capable of mending hundreds of single- and double-stranded DNA breaks without lethality or induced mutation. After the release of genome sequence of D. radiodurans wild type strain R1 in 1999, the structure characteristics,molecular defense systems and the biochemical mechanism of many key repair genes,such as recA,ssb,pprI,pprA are making great progress. But the accurate regulatory network of DNA repair in Deinococcus radiodurans is still remaining unkown in large part.In Escherichia coli, RecBCD is responsible for the repair of most of double-strand DNA damages. However, Deinococcus radiodurans doesn't have an intact RecBCD complex and its genome only encodes a RecD-like protein. In this thesis, we did a primarily bioinformatic analyses of D. radiodurans RecD and found its new role in antioxidant process.Based on the results of amino acid alignment, we found D. radiodurans RecD had a high similarity to E. coli RecD. It belongs to helicase superfamily I and has seven conserved motifs. It is composed of two domains: helicase domain and N-terminal domain.Previous studies showed that D. radiodurans RecD is a 5'-3'helicase but it acitivity is very slow. In order to study other possible functions of D. radioudurans RecD, we constructed a D. radiodurans recD disruptant using insertional mutagenesis. Phenotype analyses showed that mutation of D. radiodurans recD didn't have apparent effect on the resistance of D. radiodurans to gamma radiation and ultraviolet but increases dramatically the sensitivity of D. radiodurans to hydrogen peroxide. Complementation experiments showed that intact D. radiodurans RecD and co-expression of both helicase domain and N-terminal domain of D. radiodurans RecD could restore completely the resistance of the recD mutant to hydrogen peroxide, however, E. coli RecD, individual helicase domain or N-terminal domain of D. radiodurans RecD didn't compensate the resistance of the recD mutant to hydrogen peroxide. Further studies showed that D. radiodurans RecD participated in the antioxidant process via regulating the activities of reactive oxygen species scavenging and catalase.