title: Characterization of DRA0282 from Deinococcus radiodurans for its role in bacterial resistance to DNA damage.
abstract: DRA0282, a hypothetical protein, was found in a pool of nucleotide-binding proteins in Deinococcus radiodurans cells recovering from gamma radiation stress. This pool exhibited an unusual inhibition of nuclease activity by ATP. The N terminus of DRA0282 showed similarity to human Ku80 homologues, while the C terminus showed no similarities to known proteins. The recombinant protein required Mn(2+) for its interaction with DNA and protected dsDNA from exonuclease III degradation. The binding of the protein to supercoiled DNA with a K(d) of ~2.93 nM was nearly 20-fold stronger than its binding to ssDNA and nearly 67-fold stronger than its binding to linear dsDNA. Escherichia coli cells expressing DRA0282 showed a RecA-dependent enhancement of UV and gamma radiation tolerance. The ΔdrA0282 mutant of D. radiodurans showed a dose-dependent response to gamma radiation. At 14 kGy, the ΔdrA0282 mutant showed nearly 10-fold less survival, while at this dose both pprA : : catΔdrA0282 and pprA : : cat mutants were nearly 100-fold more sensitive than the wild-type. These results suggested that DRA0282 is a DNA-binding protein with a preference for superhelical DNA, and that it plays a role in bacterial resistance to DNA damage through a pathway in which PprA perhaps plays a dominant role in D. radiodurans.
authors.affiliation: Molecular Biology Division, Bhabha Atomic Research Centre, Mumbai 400 085, India.
authors.forename: Anubrata D
authors.initials: AD
authors.lastname: Das
authors.affiliation: Molecular Biology Division, Bhabha Atomic Research Centre, Mumbai 400 085, India.
authors.forename: Hari S
authors.initials: HS
authors.lastname: Misra
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002413:Cations, Divalent / Q000378:metabolism; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D034301:Deinococcus / Q000235:genetics / Q000378:metabolism* / Q000528:radiation effects*; D004926:Escherichia coli / Q000235:genetics / Q000528:radiation effects; D017353:Gene Deletion; D015870:Gene Expression; D007700:Kinetics; D008345:Manganese / Q000378:metabolism; D050296:Microbial Viability / Q000528:radiation effects; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D002413:Cations, Divalent; D003067:Coenzymes; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D008345:Manganese; D004247:DNA