Cloning and expression of cDNA for a human enzyme that hydrolyzes 8-oxo-dGTP, a mutagenic substrate for DNA synthesis.

Abstract

8-Oxoguanine (8-oxo-7, 8-dihydroguanine) is produced in DNA, as well as in nucleotide pools of cells, by active oxygen species normally formed during cellular metabolic processes. 8-Oxoguanine nucleotide can pair with cytosine and adenine nucleotides at almost equal efficiencies, and transversion mutation ensues. Human cells contain enzyme activity, which hydrolyzes 8-oxo-dGTP to 8-oxo-dGMP, and this enzyme is responsible for preventing misincorporation of 8-oxoguanine into DNA. We purified this particular human enzyme to physical homogeneity and determined a partial amino acid sequence. We then cloned the cDNA for human 8-oxo-dGTPase and examined its nucleotide sequence. The human protein comprises 156 amino acid residues and has some sequence homology with the Escherichia coli MutT protein, which has a distinct 8-oxo-dGTPase activity. When the human cDNA was expressed in E. coli mutT- mutant cells, there was a significant amount of 8-oxo-dGTPase activity. In such cells, the frequency of spontaneous mutation was greatly reduced. We propose that the human 8-oxo-dGTPase protects genetic information from the untoward effects of endogenous oxygen radicals.