The Effect of Tautomeric Constant on the Specificity of Nucleotide Incorporation during DNA Replication: Support for the Rare Tautomer Hypothesis of Substitution Mutagenesis
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- 作者：Harris ; Victoria H. ; Smith ; Clifford L. ; Jonathan Cummins ; W. ; Hamilton ; Alan L. ; Adams ; Harry ; et. al.
- 关键词：pyrimidine analogue ; dP ; tautomerisation ; substitution mutagenesis ; DNA replication ; dFTP ; 2 ; 4-difluorotoluene-2&prime ; -deoxyriboside-5&prime ; -triphosphate
- 刊名：Journal of Molecular Biology
- 出版年：2003
- 出版时间：March 7, 2003
- 年：2003
- 卷：326
- 期：5
- 页码：1389-1401
- 全文大小：626 K
文摘
The nucleoside analogue dP (6-(2-deoxy-β-d-ribofuranosyl)-3,4-dihydro-6H,8H-pyrimido[4,5-c][1,2]oxazin-2-one) displays ambivalent hydrogen bonding characteristics whereby the imino tautomer of P can base-pair with adenine and its amino tautomer can base-pair with guanine. Fixed imino and amino tautomers of 6-methyl-3,4-dihydro-6H,8H-pyrimido[4,5-c][1,2]oxazin-2-one (N-methyl P) have been synthesised and their structures obtained by X-ray crystallography. The tautomeric constant of N-methyl P has been calculated from pKa values of the fixed tautomers and the kinetic parameters for the incorporation of its 5′-triphosphate (dPTP) by exonuclease-free Klenow fragment of DNA polymerase I have been determined. A strong correlation between the tautomeric constant and the incorporation specificity of dPTP is found. These results lend support to the proposal that the minor tautomeric forms of the natural bases may play an important role in substitution mutagenesis during DNA replication. Furthermore, they imply that DNA polymerases impose specific steric requirements on the base-pair during nucleotide incorporation.