Intracellular Localization of 8-Oxo-dGTPase in Human Cells, with Special Reference to the Role of the Enzyme in Mitochondria (*)
- Dongchon Kang(1)(§),
- Jun-ichi Nishida(2),
- Akihiro Iyama(2),
- Yusaku Nakabeppu(2),
- Masato Furuichi(2),
- Toshiyuki Fujiwara(3),
- Mutsuo Sekiguchi(2) and
- Koichiro Takeshige(1)
- From the (1) Department of Biochemistry, Kyushu University School of Medicine, Fukuoka 812, the
- (2)Medical Institute of Bioregulation, Kyushu University, Fukuoka 812, and the
- (3)Department of Biochemistry, Fukuoka University School of Medicine, Fukuoka 814-01, Japan
- § To whom correspondence should be addressed: Dept. of Biochemistry, Kyushu University School of Medicine, 3-1-1 Maidashi, Fukuoka 812, Japan. Fax: 81-92-632-2373.
Abstract
We examined the intracellular distribution of 8-oxo-dGTPase (8-oxo-7,8-dihydrodeoxyguanosine triphosphatase) encoded by the MTH1 gene, a human mutator homologue. The activity of 8-oxo-dGTPase mainly located in cytosolic and mitochondrial soluble fractions of Jurkat cells, a human T-cell leukemia line. Electron microscopic immunocytochemistry, using a specific antibody against MTH1 protein, showed localization of MTH1 protein in the mitochondrial matrix. Activity in the mitochondria accounted for about 4% of the total activity. The specific activity in the mitochondrial soluble fraction (8093 units/mg protein) was as high as that in the cytosolic fraction (8111 unit/mg protein). The 8-oxo-dGTPase activities in cytosolic and mitochondrial soluble fractions co-eluted with MTH1 protein by anion-exchange chromatography, and the molecular mass of the mitochondrial MTH1 protein was much the same as that of the cytosolic MTH1 protein (about 18 kDa). HeLa cells expressing MTH1 cDNA showed an increased cytoplasmic signal together with a weak signal in the nucleus in in situ immunostaining of MTH1 protein, and the overexpressed MTH1 protein was recovered from both cytosolic and mitochondrial fractions. Thus, the 8-oxo-dGTPase encoded by MTH1 gene is localized in mitochondria and cytosol.
Footnotes
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↵* This work was supported in part by grants from the Ministry of Education, Science and Culture of Japan. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵1 The abbreviations used are:
- 8-oxoguanine
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8-oxo-7,8-dihydroguanine
- 8-oxo-dGTPase
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8-oxo-7,8-dihydrodeoxyguanosine triphosphatase
- APMSF
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(p-amidinophenyl)methanesulfonyl fluoride hydrochloride
- PAGE
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polyacrylamide gel electrophoresis
- HPLC
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high performance liquid chromatography.
