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ROLES OF URACIL‐DNA GLYCOSYLASE AND APYRIMIDINIC ENDONUCLEASES IN THE MOLECULAR 5‐BROMO‐2′‐DEOXYURIDINE PHOTOSENSITIZATION IN Escherichia coli K‐12
Author(s) -
Yamamoto Yoko,
Fujlwarat Yoshisada
Publication year - 1993
Publication title -
photochemistry and photobiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.818
H-Index - 131
eISSN - 1751-1097
pISSN - 0031-8655
DOI - 10.1111/j.1751-1097.1993.tb04905.x
Subject(s) - dna glycosylase , uracil dna glycosylase , uracil , microbiology and biotechnology , wild type , chemistry , dna , mutant , base excision repair , dna repair , deoxyuridine , thymine , thymidine , ap site , biology , biochemistry , gene
The molecular mechanism for 5‐bromo‐2′‐deoxyuridine (BrdU) photosensitization was studied in thymine‐requiring wild‐type and uracil‐DNA glycosylase (UDG)‐deficient ung mutant cells of Escherichia coli K‐12. Wild‐type cells were more sensitive to BrdU photosensitiLation than ung mutant cells. IJV induced the identica/ numbers of alkaline sucrose single‐strand breaks (SSB) in 5‐bromouracil‐DNA (BrU‐DNA) of both the wild type and ung mutant. The ung mutant cells repaired SSB almost completely, whereas the wild‐type cells with UDG produced more adverse SSB by 90 min after UV. Neutral agarose gel electrophoresis of minipreps indicated that UV induced (1) more smears of host BrU‐DNA possibly by more double‐strand breaks (DSB) and (2) a greater decline of pBR322 Form I BrU‐DNA in the wild‐type cells than the ung cells. These results indicated a greater induction of SSB by apyrimidinic (AP) endonucleases in wild‐type cells. The ung/ wild ratios (=1.7–1.9) for cellular and plasmid BrdU sensitizations aftcr growth in 50% BrdU were similar. The extents of UDG‐dependent and UDG‐independent sensitizations in wild‐type cells were ∼40 and ∼60%, respectively. The xth nfo double mutant defective in both exonuclcase III and endonucleasc IV was more sensitive to BrdU photosensitization than the wild type, indicating that an excess of AP sites remaining after uracil excision in the xth nfo mutant causes a greater BrdU photosensitization than SSB by AP endonucleases in wild‐type cells. Conversely, the xth rfo ung triple mutant was more resistant to BrdU photosensitization than the xth nfo double mutant, so that UV‐induced uracil residues in the BrU‐DNA are tolerated and do not appear to be directly responsible for BrdU photosensitization.