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

Yakushiji, Hiroyuki; Maraboeuf, Fabrice; Takahashi, Masayuki; Deng, Zeng Sui; Kawabata, Shun Ichiro; Nakabeppu, Yusaku; Sekiguchi, Mutsuo

doi:10.1016/s0921-8777(97)00025-6

Cited by 48 publications

(32 citation statements)

References 58 publications

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“…As shown in Table VII, mutation rates of CC101T cells harboring negative mutants used for the second mutagenesis as templates were as high as that of the cells with vector pTT100. Expression levels of the negative mutant proteins were lower than that of wild type MTH1, and their 8-oxo-dGTPase activity was detectable in only a few cases (Val 83 ) were predicted using PHD methods (27,36). In the chimeric protein, MTH1-Ec, the 23-residue sequence, which consists of loop I and ␣-helix I in MTH1 protein, is replaced with that of E. coli MutT protein (shaded), as described under "Experimental Procedures."…”

Section: ϫ6mentioning

confidence: 99%

“…Plasmid pTT100 is a derivative of pTrc99A lacking the lacI q gene. The wild type MTH1 cDNA encoding 18 kDa of Val 83 -MTH1d polypeptide was subcloned into NcoI-BamHI sites of pTT100 yielding pTT100:hMTH1 (13,27). Plasmid pALTER:MTH-ss was constructed by inserting a 327-base pair SspI-SacI fragment, encoding the conserved amino acid sequence of hMTH1 protein, into the SmaI-SacI site of pALTER-1 (28).…”

Section: Chemicals-[␣-mentioning

confidence: 99%

“…Western Blotting-Bacterial cell crude extracts, prepared as described (27), were subjected to 15% SDS-polyacrylamide gel electrophoresis and then to Western blotting. Purified 18-kDa Val 83 -MTH1d polypeptide (27) was used as a standard for quantification of mutant MTH1 protein in the bacterial extracts.…”

Section: Chemicals-[␣-mentioning

confidence: 99%

“…Purified 18-kDa Val 83 -MTH1d polypeptide (27) was used as a standard for quantification of mutant MTH1 protein in the bacterial extracts. Western blotting was done as described (28), using anti-MTH1 or anti-M77 antibodies.…”

Section: Chemicals-[␣-mentioning

confidence: 99%

“…2B, and the specific activity of MTH1 or MTH1-Ec was calculated based on their contents in crude (27). On the other hand, the specific activity of 8-oxo-dGTPase for MTH1-Ec protein was 62.5 ϫ 10 2 units/g at 37°C, that is about 1 ⁄4 that for MTH1, indicating that total 8-oxo-dGTPase activity in cells expressing MTH1-Ec at 37°C is less than 1 ⁄12 that in cells expressing MTH1 itself.…”

Section: ϫ6mentioning

confidence: 99%

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Functional Significance of the Conserved Residues for the 23-Residue Module among MTH1 and MutT Family Proteins

Fujii¹,

Shimokawa²,

Sekiguchi³

et al. 1999

Journal of Biological Chemistry

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Section: ϫ6mentioning

confidence: 99%

Section: Chemicals-[␣-mentioning

confidence: 99%

Section: Chemicals-[␣-mentioning

confidence: 99%

Section: Chemicals-[␣-mentioning

confidence: 99%

Section: ϫ6mentioning

confidence: 99%

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Functional Significance of the Conserved Residues for the 23-Residue Module among MTH1 and MutT Family Proteins

Fujii¹,

Shimokawa²,

Sekiguchi³

et al. 1999

Journal of Biological Chemistry

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Oxidative damage in nucleic acids and Parkinson's disease

Nakabeppu

Tsuchimoto

Yamaguchi

et al. 2007

J of Neuroscience Research

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263

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Oxidative DNA lesions, such as 8-oxoguanine (8-oxoG), accumulate in nuclear and mitochondrial genomes during aging, and such accumulation can increase dramatically in patients with Parkinson's disease (PD). To counteract oxidative damage to nucleic acids, human and rodents are equipped with three distinct enzymes. One of these, MTH1, hydrolyzes oxidized purine nucleoside triphosphates, such as 8-oxo-2'-deoxyguanosine triphosphate and 2-hydroxy-2'-deoxyadenosine triphosphate, to their monophosphate forms. The other two enzymes are 8-oxoG DNA glycosylase encoded by the OGG1 gene and adenine/2-hydroxyadenine DNA glycosylase encoded by the MUTYH gene. We have shown a significant increase in 8-oxoG in mitochondrial DNA as well as an elevated expression of MTH1, OGG1, and MUTYH in nigrostriatal dopaminergic neurons of PD patients, suggesting that the buildup of these lesions may cause dopamine neuron loss. We established MTH1-null mice and found that MTH1-null fibroblasts were highly susceptible to cell death caused by H(2)O(2) characterized by pyknosis and electron-dense deposits in the mitochondria, and that this was accompanied by an ongoing accumulation of 8-oxoG in nuclear and mitochondrial DNA. We also showed that MTH1-null mice exhibited an increased accumulation of 8-oxoG in striatal mitochondrial DNA, followed by more extreme neuronal dysfunction after 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine administration than that of wild-type mice. In conclusion, oxidative damage in nucleic acids is likely to be a major risk factor for Parkinson's disease, indicating that a solid understanding of the defense mechanisms involved will enable us to develop new strategies for protecting the brain against oxidative stress.

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