Purification and characterization of a novel DNA repair enzyme from the extremely radioresistant bacterium Rubrobacter radiotolerans.
Rubrobacter radiotolerans is an extremely radioresistant bacterium. It exhibits higher resistance than the well-known radioresistant bacterium Deinococcus radiodurans, but the molecular mechanisms responsible for the radio-resistance of R. radiotolerans remain unknown. In the present study, we have demonstrated the presence of a novel DNA repair enzyme in R. radiotolerans cells that recognizes radiation-induced DNA damages such as thymine glycol, urea residues, and abasic sites. The enzyme was purified from the crude cell extract by a series of chromatography to an apparent physical homogeneity. The purified enzyme showed a single band with a molecular mass of approximately 40 kDa in SDS-polyacrylamide gel electrophoresis, and was designated as R-endonuclease. R-Endonuclease exhibited repair activity for thymine glycol, urea residues, and abasic sites present in plasmid DNA, but did not act on intact DNA, UV-irradiated DNA and DNA containing reduced abasic sites. The substrate specificity together with the salt and pH optima suggests that R-endonuclease is a functional homolog of endonuclease III of Escherichia coli.
Asgarani
E
E
Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Higashi, Japan.
Terato
H
H
Asagoshi
K
K
Shahmohammadi
H R
HR
Ohyama
Y
Y
Saito
T
T
Yamamoto
O
O
Ide
H
H
D001080:Apurinic Acid / Q000032:analysis; D001173:Arthrobacter / Q000201:enzymology* / Q000235:genetics / Q000528:radiation effects; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification*; D004249:DNA Damage; D011088:DNA Ligases / Q000235:genetics / Q000302:isolation & purification*; D004260:DNA Repair / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics* / Q000528:radiation effects; D043224:Deoxyribonuclease (Pyrimidine Dimer); D004591:Electrophoresis, Polyacrylamide Gel; D004706:Endodeoxyribonucleases / Q000378:metabolism; D004720:Endonucleases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D011836:Radiation Tolerance / Q000235:genetics*; D011886:Radon; D013379:Substrate Specificity; D013941:Thymine / Q000031:analogs & derivatives / Q000378:metabolism; D014508:Urea / Q000378:metabolism; D014871:Water Microbiology
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D001080:Apurinic Acid; C029389:thymine glycol; D014508:Urea; D004706:Endodeoxyribonucleases; D004720:Endonucleases; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli; D011088:DNA Ligases; D011886:Radon; D013941:Thymine