Repair of propanodeoxyguanosine by nucleotide excision repair in vivo and in vitro.
Repair of the exocyclic DNA adduct propanodeoxyguanosine (PdG) was assessed in both in vivo and in vitro assays. PdG was site-specifically incorporated at position 6256 of M13MB102 DNA, and the adducted viral genome was electroporated into repair-proficient and repair-deficient Escherichia coli strains. Comparable frequencies of PdG --> T and PdG --> A mutations at position 6256 were detected following replication of the adducted genomes in wild-type E. coli strains. A 4-fold increase in the frequencies of transversions and transitions was observed in E. coli strains deficient in Uvr(A)BC-dependent nucleotide excision repair. A similar increase in the replication of the adduct containing strand was observed in the repair-deficient strains. No change in the frequency of targeted mutations was observed in strains deficient in one or both of the genes coding for 3-methyladenine glycosylase. Incubation of purified E. coli Uvr(A)BC proteins with a duplex 156-mer containing a single PdG adduct resulted in removal of a 12-base oligonucleotide containing the adduct. Incubation of the same adducted duplex with Chinese hamster ovary cell-free extracts also resulted in removal of the adduct. PdG was a better substrate for repair by the mammalian nucleotide excision repair complex than the bacterial repair complex and was approximately equal to a thymine-thymine dimer as a substrate for the former. The results of these in vivo and in vitro experiments indicate that PdG, a homolog of several endogenously produced DNA adducts, is repaired by the nucleotide excision repair pathway.
Johnson
K A
KA
Department of Biochemistry, Center in Molecular Toxicology and the Vanderbilt Cancer Center, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
Fink
S P
SP
Marnett
L J
LJ
D000818:Animals; D017104:Bacteriophage M13 / Q000235:genetics; D016466:CHO Cells; D002474:Cell-Free System; D006224:Cricetinae; D018736:DNA Adducts / Q000378:metabolism*; D004260:DNA Repair; D004261:DNA Replication; D004279:DNA, Viral / Q000378:metabolism; D003849:Deoxyguanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism; D029968:Escherichia coli Proteins
D018736:DNA Adducts; D004279:DNA, Viral; D029968:Escherichia coli Proteins; C060195:cyclic 1,N(2)-propanodeoxyguanosine; D004706:Endodeoxyribonucleases; C036385:endodeoxyribonuclease uvrABC; D003849:Deoxyguanosine