The Oxidized Forms of dATP Are Substrates for the Human MutT Homologue, the hMTH1 Protein

doi:10.1074/jbc.274.26.18201

The Oxidized Forms of dATP Are Substrates for the Human MutT Homologue, the hMTH1 Protein*

From the ^‡Department of Environmental Oncology, Institute of Industrial Ecological Sciences, University of Occupational and Environmental Health, 1-1 Iseigaoka, Yahatanishi-ku, Kitakyushu 807-8555, ^¶CREST, Japan Science and Technology Corporation, and the ^§Department of Biochemistry, Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan

Abstract

The possibility that Escherichia coliMutT and human MTH1 (hMTH1) hydrolyze oxidized DNA precursors other than 8-hydroxy-dGTP (8-OH-dGTP) was investigated. We report here that hMTH1 hydrolyzed 2-hydroxy-dATP (2-OH-dATP) and 8-hydroxy-dATP (8-OH-dATP), oxidized forms of dATP, but not (R)-8,5′-cyclo-dATP, 5-hydroxy-dCTP, and 5-formyl-dUTP. The kinetic parameters indicated that 2-OH-dATP was hydrolyzed more efficiently and with higher affinity than 8-OH-dGTP. 8-OH-dATP was hydrolyzed as efficiently as 8-OH-dGTP. The preferential hydrolysis of 2-OH-dATP over 8-OH-dGTP was observed at all of the pH values tested (pH 7.2 to pH 8.8). In particular, a 5-fold difference in the hydrolysis efficiencies for 2-OH-dATP over 8-OH-dGTP was found at pH 7.2. However, E. coli MutT had no hydrolysis activity for either 2-OH-dATP or 8-OH-dATP. Thus, E. coli MutT is an imperfect counterpart for hMTH1. Furthermore, we found that 2-hydroxy-dADP and 8-hydroxy-dGDP competitively inhibited both the 2-OH-dATP hydrolase and 8-OH-dGTP hydrolase activities of hMTH1. The inhibitory effects of 2-hydroxy-dADP were 3-fold stronger than those of 8-hydroxy-dGDP. These results suggest that the three damaged nucleotides share the same recognition site of hMTH1 and that it is a more important sanitization enzyme than expected thus far.

Footnotes

↵* This work was supported in part by Grants-in-aid for Scientific Research on Priority Areas from the Ministry of Education, Science, Sports and Culture of Japan.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵‖ To whom correspondence should be addressed. Tel.: 81-93-691-7468; Fax: 81-93-601-2199; E-mail:h-kasai{at}med.uoeh-u.ac.jp.
↵2 H. Kamiya and H. Kasai, unpublished results.
↵3 H. Oda, A. Taketomi, R. Maruyama, R. Itoh, K. Nishioka, H. Yakushiji, T. Suzuki, M. Sekiguchi, and Y. Nakabeppu, unpublished results.
Abbreviations:

hMTH1

human MTH1

2-OH-dATP

2-hydroxy-dATP

2-OH-dADP

2-hydroxy-dADP

8-OH-dGTP

8-hydroxy-dGTP

8-OH-dGDP

8-hydroxy-dGDP

8-OH-dATP

8-hydroxy-dATP

HPLC

high pressure liquid chromatography

8-OH-dGTPase

8-hydroxydeoxyguanosine triphosphatase

2-OH-dATPase

2-hydroxydeoxyadenosine triphosphatase
- Received April 1, 1999.
The American Society for Biochemistry and Molecular Biology, Inc.