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Molecular and biochemical characterization of Eimeria tenella hexokinase

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Abstract

Hexokinase (HK) is one of the key enzymes in the glycolytic pathway that catalyzes the phosphorylation of glucose. In the present study, we cloned the HK gene from the coccidian Eimeria tenella (EtHK), expressed EtHK as a His-tagged fusion protein, and characterized its primary biochemical features. Mutagenesis confirmed that residues S159, N216, and D217 are essential or important to the EtHK catalytic activity. EtHK exhibited high affinity for d-glucose (Km = 0.67 to 0.79 mM), but was also able to utilize 2-deoxy-d-glucose (Km = 5.66 mM), d-fructose (Km = 13.76 mM), and d-mannose (Km = 25.41 mM). We also observed that quercetin and mangiferin could inhibit the EtHK enzyme activity (IC50 values = 6.52 and 85.82 μM, respectively). Among the two inhibitors, mangiferin also inhibited the growth of E. tenella in vitro (MIC50 = 0.12 μM). These observations suggest that EtHK may be explored as potential drug target, and mangiferin and its analogs may be explored for developing anti-coccidial therapeutics.

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Acknowledgments

This work was supported in part by NSFC grants (nos. 31302087 and 31402186), NSF grant of Guangdong province (no. 2015A030313561), the Science and Technology Planning Project of Guangdong province (nos. 2014A020208057, 2014B090901035, 2015B050501007, 2014B070706011, and 2012A020100001). E. tenella hexokinase (EtHK)cDNA sequence data was obtained from the ToxoDB database under the EuPathDB (http://www.EupathDB.org).

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    Authors and Affiliations

    1. Institute of Animal Health, Guangdong Academy of Agricultural Sciences, Guangzhou, Guangdong Province, 510640 People’s Republic of China

      Mingfei Sun, Shenquan Liao, Caiyan Wu, Nanshan Qi, Minna Lv, Juan Li, Xuhui Lin, Jianfei Zhang, Mingquan Xie & Jianping Cai

    2. Open Laboratory of Veterinary Public Health and Key Laboratory of Livestock Disease Prevention of Guangdong Province, Guangzhou, 510640, People’s Republic of China

      Mingfei Sun, Shenquan Liao, Caiyan Wu, Nanshan Qi, Minna Lv, Juan Li, Xuhui Lin, Jianfei Zhang, Mingquan Xie & Jianping Cai

    3. College of Animal Science and Veterinary Medicine, Henan Agricultural University, Zhengzhou, Henan Province, 450002, People’s Republic of China

      Longxian Zhang

    4. Department of Veterinary Pathobiology, College of Veterinary Medicine & Biomedical Sciences, Texas A&M University, College Station, 77843-4467, Texas, USA

      Guan Zhu

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    Mingfei Sun and Shenquan Liao contributed equally to this work.

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    Sun, M., Liao, S., Zhang, L. et al. Molecular and biochemical characterization of Eimeria tenella hexokinase. Parasitol Res 115, 3425–3433 (2016). https://doi.org/10.1007/s00436-016-5104-4

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