Human RAD2 homolog 1 5'- to 3'-exo/endonuclease can efficiently excise a displaced DNA fragment containing a 5'-terminal abasic lesion by endonuclease activity.
Repair of abasic lesions, one of the most common types of damage found in DNA, is crucial to an organism's well-being. Studies in vitro indicate that after apurinic-apyrimidinic endonuclease cleaves immediately upstream of a baseless site, removal of the 5'-terminal sugar-phosphate residue is achieved by deoxyribophosphodiesterase activity, an enzyme-mediated beta-elimination reaction, or by endonucleolytic cleavage downstream of the baseless sugar. Synthesis and ligation complete repair. Eukaryotic RAD2 homolog 1 (RTH1) nuclease, by genetic and biochemical evidence, is involved in repair of modified DNA. Efficient endonucleolytic cleavage by RTH1 nuclease has been demonstrated for annealed primers that have unannealed 5'-tails. In vivo, such substrate structures could result from repair-related strand displacement synthesis. Using 5'-tailed substrates, we examined the ability of human RTH1 nuclease to efficiently remove 5'-terminal abasic residues. A series of upstream primers were used to increasingly displace an otherwise annealed downstream primer containing a 5'-terminal deoxyribose-5-phosphate. Until displacement of the first annealed nucleotide, substrates resisted cleavage. With further displacement, efficient cleavage occurred at the 3'-end of the tail. Therefore, in combination with strand displacement activity, RTH1 nucleases may serve as an important alternative to other pathways in repair of abasic sites in DNA.
DeMott
M S
MS
Department of Biochemistry, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA.
Shen
B
B
Park
M S
MS
Bambara
R A
RA
Zigman
S
S
D004247:DNA / Q000378:metabolism*; D053938:DNA Fragmentation; D004268:DNA-Binding Proteins; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D029701:Saccharomyces cerevisiae Proteins; D013379:Substrate Specificity
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; C073998:RAD2 protein, S cerevisiae; D004247:DNA; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases