5-Hydroxypyrimidine deoxynucleoside triphosphates are more efficiently incorporated into DNA by exonuclease-free Klenow fragment than 8-oxopurine deoxynucleoside triphosphates

Abstract

Recent studies with 8-oxodeoxyguanosine triphosphate (8-oxodGTP) have suggested that incorporation of oxidized nucleotides from the precursor pool into DNA may have deleterious effects. Here we show that 5-hydroxydeoxycytosine triphosphate (5-OHdCTP) and 5-hydroxydeoxyuridine triphosphate (5-OHdUTP) are more efficient substrates than 8-oxodGTP for Escherichia coli DNA polymerase I Klenow fragment lacking proofreading activity, while 8-oxodeoxyadenosine triphosphate (8-oxodGTP, 5-OHdCTP can mispair with dA in DNA but with lower efficiency. Since the 5-hydroxypyrimidines are present in normal and oxidized cellular DNA in amounts similar to the 8-oxopurines, these data suggest that enzymatic mechanisms might exist for removing them from the DNA precursor pools.

Identification and Characterization of a Novel Human DNA Glycosylase for Repair of Cytosine-derived Lesions

“…In any case, 5-OHU is quite abundant, and its level in DNA from mammalian tissues and human cells is comparable to that of 8-oxoguanine (20). 5-OHU pairs with A during DNA replication by Klenow polymerase, which will cause GC 3 AT transition (21). In contrast, the mammalian bypass DNA polymerase Pol preferentially incorporates T and to a lesser extent G opposite 5-OHU (19).…”

“…In contrast, only the correct nucleotide G is inserted opposite 5-OHC (19). Furthermore, oxidatively modified deoxynucleotides could be incorporated into DNA from the nucleotide pool, and 5-OHdUTP is an efficient substrate for Klenow polymerase (21).…”

Mutagenicities of 8-Hydroxyguanine and 2-Hydroxyadenine Produced by Reactive Oxygen Species

“…These explanations agree with results obtained by in vitro DNA synthesis experiments. 4,36,[40][41][42][43] As discussed in the previous paper, 47) the DNA pol-specific mispairing properties of 2-OH-dATP may be derived from the presence of the hydrophobicity-dependent enol-keto equilibrium of 2-OH-Ade, and from the possibility of adopting the syn conformation.…”

“…36,40,41) The ratio of the incorporation of 8-OH-dGTP opposite A to that opposite C depends upon the DNA pol. Misincorporation of 8-OH-dGTP opposite G or T has not been reported.…”

Mutagenic effects of 8-hydroxy-dGTP in live mammalian cells

“…DNA polymerases incorporate 8-OH-dGTP opposite C and A in vitro [7,[16][17][18][19][20][21][22][23]. In terms of the accumulation of 8-OH-Gua in DNA, the contribution of 8-OH-dGTP from the nucleotide pool and the direct oxidation of G bases in DNA has been reported to be almost equal [24].…”