Recognition and incision of site-specifically modified C8 guanine adducts formed by 2-aminofluorene, N-acetyl-2-aminofluorene and 1-nitropyrene by UvrABC nuclease.
Nucleotide excision repair plays a crucial role in removing many types of DNA adducts formed by UV light and chemical carcinogens. We have examined the interactions of Escherichia coli UvrABC nuclease proteins with three site-specific C8 guanine adducts formed by the carcinogens 2-aminofluorene (AF), N:-acetyl-2-acetylaminofluorene (AAF) and 1-nitropyrene (1-NP) in a 50mer oligonucleotide. Similar to the AF and AAF adducts, the 1-NP-induced DNA adduct contains an aminopyrene (AP) moiety covalently linked to the C8 position of guanine. The dissociation constants for UvrA binding to AF-, AAF- and AP-DNA adducts, determined by gel mobility shift assay, are 33 +/- 9, 8 +/- 2 and 23 +/- 9 nM, respectively, indicating that the AAF adduct is recognized much more efficiently than the other two. Incision by UvrABC nuclease showed that AAF-DNA was cleaved approximately 2-fold more efficiently than AF- or AP-DNA (AAF > AF approximately AP), even though AP has the largest molecular size in this group. However, an opened DNA structure of six bases around the adduct increased the incision efficiency for AF-DNA (but not for AP-DNA), making it equivalent to that for AAF-DNA. These results are consistent with a model in which DNA damage recognition by the E. coli nucleotide excision repair system consists of two sequential steps. It includes recognition of helical distortion in duplex DNA followed by recognition of the type of nucleotide chemical modification in a single-stranded region. The difference in incision efficiency between AF- and AAF-DNA adducts in normal DNA sequence, therefore, is a consequence of their difference in inducing structural distortions in DNA. The results of this study are discussed in the light of NMR solution structures of these DNA adducts.
Luo
C
C
Sealy Center for Molecular Science and Department of Human Biological Chemistry and Genetics, University of Texas Medical Branch, Galveston, TX 77555, USA.
Krishnasamy
R
R
Basu
A K
AK
Zou
Y
Y
D015073:2-Acetylaminofluorene / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D001483:Base Sequence; D018736:DNA Adducts / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004249:DNA Damage / Q000235:genetics; D004260:DNA Repair / Q000235:genetics; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D005449:Fluorenes / Q000737:chemistry / Q000378:metabolism; D006147:Guanine / Q000031:analogs & derivatives* / Q000378:metabolism*; D007700:Kinetics; D009153:Mutagens / Q000737:chemistry / Q000378:metabolism*; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011721:Pyrenes / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity; D013816:Thermodynamics
D018736:DNA Adducts; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D005449:Fluorenes; D009153:Mutagens; D009838:Oligodeoxyribonucleotides; D011721:Pyrenes; C012177:2-aminofluorene; D006147:Guanine; D015073:2-Acetylaminofluorene; D004706:Endodeoxyribonucleases; C036385:endodeoxyribonuclease uvrABC; C032668:1-nitropyrene