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Molecular identification, immunolocalization, and characterization of Clonorchis sinensis triosephosphate isomerase

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Abstract

Clonorchis sinensis triosephosphate isomerase (CsTIM) is a key regulatory enzyme of glycolysis and gluconeogenesis, which catalyzes the interconversion of glyceraldehyde 3-phosphate to dihydroxyacetone phosphate. In this study, the biochemical characterizations of CsTIM have been examined. A full-length complementary DNA (cDNA; Cs105350) sequence encoding CsTIM was obtained from our C. sinensis cDNA library. The open reading frame of CsTIM contains 759 bp which encodes 252 amino acids. The amino acid sequence of CsTIM shares 60–65 % identity with other species. Western blot analysis displayed that recombinant CsTIM (rCsTIM) can be probed by anti-rCsTIM rat serum and anti-C. sinensis excretory/secretory products (anti-CsESPs) rat serum. Quantitative reverse transcription (RT)-PCR and western blotting analysis revealed that CsTIM messenger RNA (mRNA) and protein were differentially expressed in development cycle stages of the parasite, including adult worm, metacercaria, excysted metacercaria, and egg. In addition, immunolocalization assay showed that CsTIM was located in the seminal vesicle, eggs, and testicle. Moreover, rCsTIM exhibited active enzyme activity in catalytic reactions. The Michaelis constant (K m) of rCsTIM was 0.33 mM, when using glyceraldehyde 3-phosphate as the substrate. The optimal temperature and pH of CsTIM were 37 °C and 7.5–9.5, respectively. Collectively, these results suggest that CsTIM is an important protein involved in glycometabolism, and CsTIM possibly take part in many biological functions in the growth and development of C. sinensis.

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Acknowledgments

This study was funded by the National Natural Science Foundation of China (No. 81101270 and No. 81171602.) and the National Important Sci-tech Special Projects (2012ZX10004220).

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    Authors and Affiliations

    1. Department of Clinical Laboratory, Children’s Hospital of Zheng Zhou, Zheng Zhou, 450053, Henan, China

      Juanjuan Zhou

    2. Microscopy Lab, Gannan Medical University, Ganzhou, 341000, Jiangxi, China

      Hua Liao

    3. Department of Pathology and Pathophysiology, Henan University of Traditional Chinese Medicine, Zhengzhou, 450000, China

      Shan Li

    4. Department of Parasitology, Zhongshan School of Medicine, Sun Yat-sen University, No 74, The Second Zhongshan RD, Guangzhou, 510080, Guangdong, China

      Chenhui Zhou, Yan Huang, Xuerong Li, Chi Liang & Xinbing Yu

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

      Chenhui Zhou, Yan Huang, Xuerong Li, Chi Liang & Xinbing Yu

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    Correspondence to Xinbing Yu.

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    Juanjuan Zhou and Hua Liao contributed equally to this work.

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    Zhou, J., Liao, H., Li, S. et al. Molecular identification, immunolocalization, and characterization of Clonorchis sinensis triosephosphate isomerase. Parasitol Res 114, 3117–3124 (2015). https://doi.org/10.1007/s00436-015-4530-z

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