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Biochemical characterization and functional analysis of fructose-1,6-bisphosphatase from Clonorchis sinensis

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Abstract

Fructose-1,6-bisphosphatase (FBPase), a key regulatory enzyme of gluconeogenesis, plays an essential role in metabolism and development of most organisms. To the wealth of available knowledge about FBPase from Clonorchis sinensis (CsFBPase), in this study, the characteristics of CsFBPase and its potential role in pathogenesis of clonorchiasis were investigated. The Km value of CsFBPase was calculated to be 41.9 uM. The optimal temperature and pH of CsFBPase were 37 °C and pH 7.5–8.0, respectively. In addition, Mg2+ or K+ played a regulatory role in enzyme activity of CsFBPase. Both transcriptional and translational level of CsFBPase were higher in metacercariae (one of larva stages) than those in adult worm (P < 0.05). CsFBPase were observed to extensively express in the intestine, vitellaria and tegument of adult worms and ubiquitously in metacercariae. Moreover, CsFBPase was confirmed as a component of excretory/secretory products. Consequently, the translocation of CsFBPase could be detected on epithelial cells of bile duct in liver of C. sinensis infected rat. Recombinant CsFBPase can specifically bind to the membrane of human hepatic stellate cell line LX-2 by immunofluorescence analysis and stimulated proliferation and activation of LX-2 which demonstrated by Cell Counting Kit-8 and upregulation of key fibrosis-related factors, such as α-smooth muscle actin, collagen I and collagen III using qRT-PCR. Thus, we predicated that CsFBPase might be a multifunctional enzyme which played as both regulatory enzyme and virulence factor in pathogenesis of C. sinensis infection.

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Acknowledgments

This work was supported by National Key Basic Research and Development Project of China (973 project; No. 2010CB530000), The National S & T Major Program (No. 2012ZX10004-220), National Natural Science Foundation of China (No. 81101270 and No. 81171602), Fundamental Research Funds for the Central Universities (No. 3164015) and China Postdoctoral Science Foundation (No. 20110490952).

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Authors and Affiliations

  1. Department of Parasitology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, 510080, China

    Pei Liang, Yan Huang, Fan Zhang, Juanjuan Zhou, Yue Hu, Xiaoyun Wang, Chi Liang, Yanquan Xu, Qiang Mao, Xuchu Hu, Xuerong Li, Jin Xu & Xinbing Yu

  2. Key Laboratory for Tropical Diseases Control, Ministry of Education, Sun Yat-sen University, Guangzhou, 510080, China

    Pei Liang, Yan Huang, Fan Zhang, Juanjuan Zhou, Yue Hu, Xiaoyun Wang, Chi Liang, Yanquan Xu, Qiang Mao, Xuchu Hu, Xuerong Li, Jin Xu & Xinbing Yu

  3. Guangdong Provincial Institute of Public Health, Guangdong Provincial Center for Disease Control and Prevention, Guangzhou, 510300, China

    Jiufeng Sun

  4. Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, 510120, Guangdong, China

    Minghui Zheng

  5. Department of Pathogen Biology, Hainan Medical College, Haikou, 571199, Hainan, China

    Gang Lu

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  1. Pei Liang
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  3. Yan Huang
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  15. Gang Lu
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Correspondence to Gang Lu or Xinbing Yu.

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Pei Liang and Jiufeng Sun contributed equally to this article.

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Liang, P., Sun, J., Huang, Y. et al. Biochemical characterization and functional analysis of fructose-1,6-bisphosphatase from Clonorchis sinensis . Mol Biol Rep 40, 4371–4382 (2013). https://doi.org/10.1007/s11033-013-2508-4

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