Mechanistic studies of the inhibition of MutT dGTPase by the carcinogenic metal Ni(II).
Promutagenic 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG) levels are increased in DNA of animals exposed to carcinogenic metals, such as Ni(II). Besides being generated directly in genomic DNA, 8-oxo-dG may be incorporated there from 8-oxo-7,8-dihydro-2'-deoxyguanosine 5'-triphosphate (8-oxo-dGTP), a product of oxidative damage to the nucleotide pool. The Escherichia coli dGTPase MutT, and analogous dGTPases in rats and humans, have been suggested as a defense against such incorporation because they hydrolyze 8-oxo-dGTP to 8-oxo-7,8-dihydro-2'-deoxyguanosine 5'-monophosphate (8-oxo-dGMP). MutT and its mammalian counterparts are Mg(II)-dependent enzymes. Ni(II), in turn, is known to interact antagonistically with Mg(II) in biological systems. Thus, we hypothesized that Ni(II) might inhibit the activity of MutT. As an initial examination of this hypothesis, we conducted enzyme kinetic studies of MutT to determine the effect of Ni(II) on MutT activity and the mechanisms involved. As found, Ni(II) inhibited MutT in a concentration-dependent manner when either dGTP or 8-oxo-dGTP was the nucleotide substrate. Ni(II) was determined to be an uncompetitive inhibitor of MutT with respect to Mg(II) when dGTP was the substrate, with apparent Ki of 1.2 mM Ni(II), and a noncompetitive inhibitor with respect to Mg(II) when 8-oxo-dGTP was the substrate, with apparent Ki of 0.9 mM Ni(II). Hence, the two metal cations did not compete with each other for binding at the MutT active site. This makes it difficult to predict Ni(II) effects on 8-oxo-dGTPases of other species. However, based upon the amino acid sequences of human and rat MutT-like dGTPases, their capacity for Ni(II) binding should be greater than that of MutT. Whether this could lead to stronger inhibition of those enzymes by Ni(II), or not, remains to be investigated.
Porter
D W
DW
Laboratory of Comparative Carcinogenesis, National Cancer Institute-FCRDC, Frederick, Maryland 21702, USA.
Nelson
V C
VC
Fivash
M J
MJ
Kasprzak
K S
KS
D000818:Animals; D002273:Carcinogens / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D004926:Escherichia coli / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008956:Models, Chemical; D009532:Nickel / Q000494:pharmacology*; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors* / Q000235:genetics; D051381:Rats
D002273:Carcinogens; D009532:Nickel; D010744:Phosphoric Monoester Hydrolases