Structure of human MTH1, a Nudix family hydrolase that selectively degrades oxidized purine nucleoside triphosphates.
Oxygen radicals generated through normal cellular respiration processes can cause mutations in genomic and mitochondrial DNA. Human MTH1 hydrolyzes oxidized purine nucleoside triphosphates, such as 8-oxo-dGTP and 2-hydroxy-dATP, to monophosphates, thereby preventing the misincorporation of these oxidized nucleotides during replication. Here we present the solution structure of MTH1 solved by multidimensional heteronuclear NMR spectroscopy. The protein adopts a fold similar to that of Escherichia coli MutT, despite the low sequence similarity between these proteins outside the conserved Nudix motif. The substrate-binding pocket of MTH1, deduced from chemical shift perturbation experiments, is located at essentially the same position as in MutT; however, a pocket-forming helix is largely displaced in MTH1 (approximately 9 A) such that the shape of the pocket differs between the two proteins. Detailed analysis of the pocket-forming residues enabled us to identify Asn33 as one of the key residues in MTH1 for discriminating the oxidized form of purine, and mutation of this residue modifies the substrate specificity. We also show that MTH1 catalyzes hydrolysis of 8-oxo-dGTP through nucleophilic substitution of water at the beta-phosphate.
Mishima
Masaki
M
Graduate School of Biological Sciences, Nara Institute of Science and Technology, 8916-5 Takayama, Ikoma, Nara 630-0101, Japan.
Sakai
Yasunari
Y
Itoh
Noriyuki
N
Kamiya
Hiroyuki
H
Furuichi
Masato
M
Takahashi
Masayuki
M
Yamagata
Yuriko
Y
Iwai
Shigenori
S
Nakabeppu
Yusaku
Y
Shirakawa
Masahiro
M
D000595:Amino Acid Sequence; D001216:Asparagine; D001665:Binding Sites; D017124:Conserved Sequence; D045643:DNA Repair Enzymes / Q000737:chemistry* / Q000378:metabolism*; D003848:Deoxyguanine Nucleotides / Q000378:metabolism; D004926:Escherichia coli / Q000737:chemistry; D029968:Escherichia coli Proteins / Q000737:chemistry; D006801:Humans; D006860:Hydrogen Bonding; D006868:Hydrolysis; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D010084:Oxidation-Reduction; D010744:Phosphoric Monoester Hydrolases / Q000737:chemistry* / Q000378:metabolism*; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011685:Purine Nucleotides / Q000378:metabolism*; D011755:Pyrophosphatases; D011994:Recombinant Proteins; D016415:Sequence Alignment; D013379:Substrate Specificity
D003848:Deoxyguanine Nucleotides; D029968:Escherichia coli Proteins; D011685:Purine Nucleotides; D011994:Recombinant Proteins; C078206:8-oxodeoxyguanosine triphosphate; D001216:Asparagine; D010744:Phosphoric Monoester Hydrolases; D011755:Pyrophosphatases; C052211:mutT protein, E coli; C078207:8-oxodGTPase; D045643:DNA Repair Enzymes