We found that hydroxylation occurs at the C-2 position of adenine by oxygen 
radical treatment (Fe2+-EDTA) of dA, dATP, and single- and double-stranded DNA. 
This oxidatively damaged base, 2-hydroxyadenine, was produced 3-6-fold and 
40-fold less than 8-hydroxyguanine when monomers and polynucleotides, 
respectively, were treated. To determine whether the damaged nucleotide, 
2-hydroxydeoxyadenosine triphosphate (2-OH-dATP), is incorporated into a growing 
DNA, and to reveal the kinds of nucleotides opposite which 2-OH-dATP is 
incorporated, calf thymus DNA polymerase alpha and the Klenow fragment of 
Escherichia coli DNA polymerase I were used in vitro DAN synthesis in the 
presence of 2-OH-dATP. DNA polymerase alpha incorporated the nucleotide opposite 
T and C in the DNA template. On the other hand, in an experiment using the 
Klenow fragment, incorporation of 2-OH-dATP was observed only opposite T. 
Steady-state kinetic studies indicated that incorporation of 2-OH-dATP by DNA 
polymerase alpha opposite T was favored over that opposite C by a factor of only 
4.5. These results indicate that 2-OH-dATP, an oxidatively damaged nucleotide, 
is a substrate for DNA polymerases and is incorporated incorrectly by the 
replicative DNA polymerase.