A Molecular Basis for the Selective Recognition of 2-Hydroxy-dATP and 8-Oxo-dGTP by Human MTH1

Abstract

MTH1 hydrolyzes oxidized purine nucleoside triphosphates such as 8-oxo-dGTP, 8-oxo-dATP, 2-hydroxydATP, and 2-hydroxy rATP to monophosphates, and thus avoids errors caused by their misincorporation during DNA replication or transcription, which may result in carcinogenesis or neurodegeneration. This substrate specificity for oxidized purine nucleoside triphosphates was investigated by mutation analyses based on the sequence comparison with the Escherichia coli homolog, MutT, which hydrolyzes only 8-oxo-dGTP and 8-oxo-rGTP but not oxidized forms of dATP or ATP. Neither a replacement of the phosphohydrolase module of MTH1 with that of MutT nor deletions of the C-terminal region of MTH1, which is unique for MTH1, altered the substrate specificity of MTH1. In contrast, the substitution of residues at position Trp-117 and Asp-119 of MTH1, which showed apparent chemical shift perturbations with 8-oxo-dGDP in NMR analyses but are not conserved in MutT, affected the substrate specificity. Trp-117 is essential for MTH1 to recognize both 8-oxodGTP and 2-hydroxy-dATP, whereas Asp-119 is only essential for recognizing 2-hydroxy-dATP, thus suggesting that origins of the substrate-binding pockets for MTH1 and MutT are different.Endogenous oxidation of nucleic acids, such as DNA, RNA, and their precursors by reactive oxygen species generated under normal metabolic conditions, appear to induce spontaneous mutagenesis and cell death, which have been implicated in aging and various diseases including cancer and neuronal degeneration (1). It has been established that 8-oxoguanine (8-oxoG), an oxidized form of guanine, is highly mutagenic and one of the main endogenous sources for spontaneous mutagenesis. During DNA replication, 8-oxo-deoxyguanosine triphosphate (8-oxo-dGTP) can be inserted into the nascent strand opposite adenine and cytosine in the template, with almost equal efficiency, thus resulting in an A:T to C:G transversion mutation (2, 3). Accumulation of 8-oxoG in double-stranded DNA is caused by direct oxidization of guanine in DNA or incorporation of precursor nucleotide, 8-oxo-dGTP, which is generated in nucleotide pools with oxidation of dGTP (4). A misincorporation of oxidized nucleotides also occurs during transcription, thus leading to an accumulation of abnormal proteins (5, 6).To eliminate such deleterious oxidized nucleotides, organisms come equipped with elaborate mechanisms. In Escherichia coli the MutT protein hydrolyzes 8-oxo-dGTP and 8-oxoguanosine triphosphate (8-oxo-rGTP) to the monophosphate form, thus avoiding the misincorporation of mutagenic nucleotides into DNA and mRNA during replication and transcription (7,8). The absence of the mutT gene increases the spontaneous occurrence of A:T to C:G transversion a thousandfold over the wild type level (7, 9), and further increases the occurrence of transcriptional errors (6). Enzymatic activities similar to that of MutT had been identified in both human and other mammalian cells (10 -13). Based on the similarity in their amino acid sequences to that of Mut...

Oxidative nucleotide damage: consequences and prevention

“…MTH1 hydrolyzes 8-oxo-dATP, 2-hydroxy-dATP and 2-hydroxyATP as well as 8-oxodGTP (Sakai et al, 2002). This is in contrast with MutT, which acts on 8-oxoguanine-containing nucleotides alone.…”

Mitochondrial dysfunction in schizophrenia: evidence for compromised brain metabolism and oxidative stress

“…In addition, 8-oxo-G, 2-hydroxy-A, 8-oxo-A, 2-hydroxy-A are some of the most common nucleotide mutations that occur as a result of oxidative stress. 52 These oxidized purines are converted to monophosphates by NUDT1 (nucleoside diphosphate-linked moiety X motif 1, also MTH1) to prevent the inclusion of the base during transcription and DNA synthesis.…”

TH588, an MTH1 inhibitor, enhances phenethyl isothiocyanate‑induced growth inhibition in pancreatic cancer cells

“…The dNTP used in DNA synthesis are up to 13,000-fold more susceptible to oxidative damage than bases in duplex DNA (9). MuT homolog-1 (MTH1) is a pyrophosphatase that acts on oxidized nucleotides and hydrolyzes 8-oxo-2'-deoxyguanosine triphosphate in the dNTP pool to prevent its incorporation into nuclear and mitochondrial DNA, which reduces cytotoxicity in cells (10,11).…”

“…The dNTP used in DNA synthesis are up to 13,000-fold more susceptible to oxidative damage than bases in duplex DNA (9). MuT homolog-1 (MTH1) is a pyrophosphatase that acts on oxidized nucleotides and hydrolyzes 8-oxo-2'-deoxyguanosine triphosphate in the dNTP pool to prevent its incorporation into nuclear and mitochondrial DNA, which reduces cytotoxicity in cells (10,11).MTH1 is overexpressed in cancer cells and prevents oxidized nucleotides from being misincorporated into DNA. Although MTH1 is non-essential in normal cells, cancer cells require MTH1 activity to avoid the incorporation of oxidized dNTPs, which would result in DNA damage and cell death (12)(13)(14).…”