Nucleic Acids Research, Vol 25, Issue 3 597-603, Copyright © 1997 by Oxford University Press
H Shimizu, R Yagi, Y Kimura, K Makino, H Terato, Y Ohyama and H Ide
alpha-2'-Deoxyadenosine (alpha) is a major adenine lesion produced by
gamma-ray irradiation of DNA under anoxic conditions. In this study,
single-stranded recombinant M13 vectors containing alpha were constructed
and transfected into Escherichia coli to assess lethal and mutagenic
effects of this lesion. The data for alpha were further compared with those
obtained with M13 vectors containing normal A or a model abasic site (F) at
the same site. The transfection assay revealed that alpha constituted a
moderate block to DNA replication. The in vivo replication capacity to pass
through alpha was approximately 20% relative to normal A, but 20-fold
higher than that of F constituting an almost absolute replication block.
Similar data were obtained by in vitro replication of oligonucleotide
templates containing alpha or F by E.coli DNA polymerase I. The mutagenic
consequence of replicating M13 DNA containing alpha was analyzed by direct
DNA sequencing of progeny phage. Mutagenesis was totally targeted at the
site of alpha introduced into the vector. Mutation was exclusively a single
nucleotide deletion and no base substitutions were detected. The deletion
frequency associated alpha was dependent on the 3'-nearest neighbor base:
with the 3'-nearest neighbor base T mutation (deletion) frequency was 26%,
whereas 1% with the 3'-nearest neighbor base G. A possible mechanism of the
single nucleotide deletion associated with alpha is discussed on the basis
of the misinsertion-strand slippage model.
ARTICLES
Replication bypass and mutagenic effect of alpha-deoxyadenosine site- specifically incorporated into single-stranded vectors
Department of Polymer Science and Engineering, Kyoto Institute of Technology, Matsugasaki, Sakyo-ku, Kyoto 606, Japan.
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