Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em>
<i>Toxoplasma gondii</i> is a protozoan parasite that causes toxoplasmosis and infects almost one-third of the global human population. A lack of effective drugs and vaccines and the emergence of drug resistant parasites highlight the need for the development of new drugs. The mitochondr...
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2021-07-01
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| Series: | International Journal of Molecular Sciences |
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| author | Rajib Acharjee Keith K. Talaam Endah D. Hartuti Yuichi Matsuo Takaya Sakura Bundutidi M. Gloria Shinya Hidano Yasutoshi Kido Mihoko Mori Kazuro Shiomi Masakazu Sekijima Tomoyoshi Nozaki Kousuke Umeda Yoshifumi Nishikawa Shinjiro Hamano Kiyoshi Kita Daniel K. Inaoka |
| author_facet | Rajib Acharjee Keith K. Talaam Endah D. Hartuti Yuichi Matsuo Takaya Sakura Bundutidi M. Gloria Shinya Hidano Yasutoshi Kido Mihoko Mori Kazuro Shiomi Masakazu Sekijima Tomoyoshi Nozaki Kousuke Umeda Yoshifumi Nishikawa Shinjiro Hamano Kiyoshi Kita Daniel K. Inaoka |
| author_sort | Rajib Acharjee |
| collection | DOAJ |
| description | <i>Toxoplasma gondii</i> is a protozoan parasite that causes toxoplasmosis and infects almost one-third of the global human population. A lack of effective drugs and vaccines and the emergence of drug resistant parasites highlight the need for the development of new drugs. The mitochondrial electron transport chain (ETC) is an essential pathway for energy metabolism and the survival of <i>T. gondii</i>. In apicomplexan parasites, malate:quinone oxidoreductase (MQO) is a monotopic membrane protein belonging to the ETC and a key member of the tricarboxylic acid cycle, and has recently been suggested to play a role in the fumarate cycle, which is required for the cytosolic purine salvage pathway. In <i>T. gondii</i>, a putative MQO (TgMQO) is expressed in tachyzoite and bradyzoite stages and is considered to be a potential drug target since its orthologue is not conserved in mammalian hosts. As a first step towards the evaluation of TgMQO as a drug target candidate, in this study, we developed a new expression system for TgMQO in FN102(DE3)TAO, a strain deficient in respiratory cytochromes and dependent on an alternative oxidase. This system allowed, for the first time, the expression and purification of a mitochondrial MQO family enzyme, which was used for steady-state kinetics and substrate specificity analyses. Ferulenol, the only known MQO inhibitor, also inhibited TgMQO at IC<sub>50</sub> of 0.822 μM, and displayed different inhibition kinetics compared to <i>Plasmodium falciparum</i> MQO. Furthermore, our analysis indicated the presence of a third binding site for ferulenol that is distinct from the ubiquinone and malate sites. |
| first_indexed | 2024-03-10T09:16:21Z |
| format | Article |
| id | doaj.art-d870a17f03504ddcb7537dcd78279046 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T09:16:21Z |
| publishDate | 2021-07-01 |
| publisher | MDPI AG |
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| series | International Journal of Molecular Sciences |
| spelling | doaj.art-d870a17f03504ddcb7537dcd782790462023-11-22T05:39:04ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-07-012215783010.3390/ijms22157830Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em>Rajib Acharjee0Keith K. Talaam1Endah D. Hartuti2Yuichi Matsuo3Takaya Sakura4Bundutidi M. Gloria5Shinya Hidano6Yasutoshi Kido7Mihoko Mori8Kazuro Shiomi9Masakazu Sekijima10Tomoyoshi Nozaki11Kousuke Umeda12Yoshifumi Nishikawa13Shinjiro Hamano14Kiyoshi Kita15Daniel K. Inaoka16Program for Nurturing Global Leaders in Tropical and Emerging Communicable Disease, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 852-8523, JapanProgram for Nurturing Global Leaders in Tropical and Emerging Communicable Disease, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 852-8523, JapanProgram for Nurturing Global Leaders in Tropical and Emerging Communicable Disease, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 852-8523, JapanSchool of Tropical Medicine and Global Health, Nagasaki University, Nagasaki 852-8523, JapanSchool of Tropical Medicine and Global Health, Nagasaki University, Nagasaki 852-8523, JapanProgram for Nurturing Global Leaders in Tropical and Emerging Communicable Disease, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 852-8523, JapanDepartment of Immune Regulation, The Research Center for Hepatitis and Immunology, National Center for Global Health and Medicine, Chiba 272-8516, JapanDepartment of Parasitology and Research Center for Infectious Disease Sciences, Graduate School of Medicine, Osaka City University, Osaka 545-8585, JapanBiological Resource Center, NITE, Kisarazu, Chiba 292-0818, JapanGraduate School of Infection Control Sciences, Kitasato University, Tokyo 108-0072, JapanDepartment of Advanced Computational Drug Discovery Unit, Tokyo Institute of Technology, Yokohama 226-8501, JapanDepartment of Biomedical Chemistry, Graduate School of Medicine, The University of Tokyo, Tokyo 113-0033, JapanPathology Division, Aquaculture Research Department, Fisheries Technology Institute, Japan Fisheries Research and Education Agency, Minamiise 516-0193, JapanResearch Unit for Host Defense, National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Obihiro 080-8555, JapanProgram for Nurturing Global Leaders in Tropical and Emerging Communicable Disease, Graduate School of Biomedical Sciences, Nagasaki University, Nagasaki 852-8523, JapanSchool of Tropical Medicine and Global Health, Nagasaki University, Nagasaki 852-8523, JapanSchool of Tropical Medicine and Global Health, Nagasaki University, Nagasaki 852-8523, Japan<i>Toxoplasma gondii</i> is a protozoan parasite that causes toxoplasmosis and infects almost one-third of the global human population. A lack of effective drugs and vaccines and the emergence of drug resistant parasites highlight the need for the development of new drugs. The mitochondrial electron transport chain (ETC) is an essential pathway for energy metabolism and the survival of <i>T. gondii</i>. In apicomplexan parasites, malate:quinone oxidoreductase (MQO) is a monotopic membrane protein belonging to the ETC and a key member of the tricarboxylic acid cycle, and has recently been suggested to play a role in the fumarate cycle, which is required for the cytosolic purine salvage pathway. In <i>T. gondii</i>, a putative MQO (TgMQO) is expressed in tachyzoite and bradyzoite stages and is considered to be a potential drug target since its orthologue is not conserved in mammalian hosts. As a first step towards the evaluation of TgMQO as a drug target candidate, in this study, we developed a new expression system for TgMQO in FN102(DE3)TAO, a strain deficient in respiratory cytochromes and dependent on an alternative oxidase. This system allowed, for the first time, the expression and purification of a mitochondrial MQO family enzyme, which was used for steady-state kinetics and substrate specificity analyses. Ferulenol, the only known MQO inhibitor, also inhibited TgMQO at IC<sub>50</sub> of 0.822 μM, and displayed different inhibition kinetics compared to <i>Plasmodium falciparum</i> MQO. Furthermore, our analysis indicated the presence of a third binding site for ferulenol that is distinct from the ubiquinone and malate sites.https://www.mdpi.com/1422-0067/22/15/7830toxoplasmosiselectron transport chainmitochondriamembrane proteinenzyme inhibitionferulenol |
| spellingShingle | Rajib Acharjee Keith K. Talaam Endah D. Hartuti Yuichi Matsuo Takaya Sakura Bundutidi M. Gloria Shinya Hidano Yasutoshi Kido Mihoko Mori Kazuro Shiomi Masakazu Sekijima Tomoyoshi Nozaki Kousuke Umeda Yoshifumi Nishikawa Shinjiro Hamano Kiyoshi Kita Daniel K. Inaoka Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em> International Journal of Molecular Sciences toxoplasmosis electron transport chain mitochondria membrane protein enzyme inhibition ferulenol |
| title | Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em> |
| title_full | Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em> |
| title_fullStr | Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em> |
| title_full_unstemmed | Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em> |
| title_short | Biochemical Studies of Mitochondrial Malate: Quinone Oxidoreductase from <em>Toxoplasma gondii</em> |
| title_sort | biochemical studies of mitochondrial malate quinone oxidoreductase from em toxoplasma gondii em |
| topic | toxoplasmosis electron transport chain mitochondria membrane protein enzyme inhibition ferulenol |
| url | https://www.mdpi.com/1422-0067/22/15/7830 |
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