Biochemical and physicochemical characterization of normal and variant forms of human MTH1 protein with antimutagenic activity

8-0xo-7,8-dihydro-2'-deoxyguanosine 5'-triphosphate (8-oxo-dGTP) is produced during cellular metabolism, and its misincorporation into DNA causes mutation. Human cells possess an enzyme that hydrolyzes 8-oxo-dGTP to the corresponding nucleoside monophosphate, thereby preventing misincorporation of 8-oxo-7,8-dihydroguanine into DNA. Sequence an analyses of the MTH1 gene, encoding the 8-oxo-7,8-dihydro-2'-deoxyguanosine 5'-triphosphatase (8-oxo-dGTPase) protein in human cell lines revealed that a G to A base substitution frequently occurs at codon 83, which causes a change of valine to methionine in the MTH1 protein [Wu, C. et al., Biochem. Biophys. Res. Commun. 214 (1995) 1239-1245]. Here we isolated cDNAs for the two types of MTH1 protein and expressed them in Escherichia coli mutT^- cells, devoid of their own 8-oxo-dGTPase activity. The two forms of proteins were purified to physical homogeneity, and amino acid analyses confirmed that the variant protein, Met⁸³-MTH1, indeed carries the corresponding amino acid substitution. Met⁸³-MTH1, but not normal type Val⁸³-MTH1, was separated into two peaks in hydrophobic interacting chromatography. 8-Oxo-dGTPase activity of Met⁸³-MTH1 is more thermolabile than that of Val⁸³-MTH1. Circular dichroism (CD) and fluorescence spectroscopic analyses confirmed this conclusion. CD further indicated that Met⁸³-MTH1 has a higher α-helix content. Substitution of valine for methionine at the residue 83 of MYH1 protein appears to lead to alteration in the secondary structure which renders the protein more labile than the normal type protein.