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Molecular cloning and characterization of a cathepsin B from Angiostrongylus cantonensis

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Abstract

Cysteine proteases, a superfamily of hydrolytic enzymes, have numerous functions in parasites. Here, we reported the cloning and characterization of a cDNA encoding a cathepsin B (AcCPB) from Angiostrongylus cantonensis fourth-stage larvae cDNA library. The deduced amino acid sequence analysis indicated AcCPB is related to other cathepsin B family members with an overall conserved architecture. AcCPB is evolutionarily more close to other parasitic nematode cathepsin B than the ones from hosts, sharing 43–53% similarities to the homologues from other organisms. Real-time quantitative PCR analysis revealed that AcCPB was expressed significantly higher in the fourth-stage larvae (L4) and the fifth-stage larvae (L5) than that in the third-stage larvae (L3) and adult worms (Aw). Unexpectedly, AcCPB was expressed at a higher level in L4 and L5 derived from mice than the larvae at the same stages derived from rats. The protease activity of recombinant AcCPB (rAcCPB) expressed in Escherichia coli showed high thermostability and acidic pH optima. The role in ovalbumin digestion and enzyme activity of rAcCPB could be evidently inhibited by cystatin from A.cantonensis. Furthermore, we found rAcCPB increased the expression levels of CD40, MHC II, and CD80 on LPS-stimulated dendritic cells (DCs). In this study, we provided the first experimental evidence for the expression of cathepsin B in A.cantonensis. Besides its highly specific expression in the stages of L4 and L5 when the worms cause dysfunction of the blood–brain barrier of hosts, AcCPB displayed different expression profiles in non-permissive host- and permissive host-derived larval stages and was involved in the maturation of DCs, suggesting a potential role in the central nervous system invasion and the immunoregulation during parasite–host interactions.

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Acknowledgments

We are grateful to Dr. Jue-heng Wu, Yue Zhu, Sen-mao Li, Li-li Zhang, and Zi-ran Zhao for their expert technical assistances in mass spectrometry, fluorescence microplate reader and assay for enzyme activity. This work was supported by grants from the National Basic Research Program of China (2010CB530004), the National Natural Science Foundation of China (30771888, 30800966), and Research Fund for Students of Sun Yat-sen University (2011).

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  1. Department of Parasitology, Zhongshan School of Medicine, Sun Yat-sen University, 74 2nd Zhongshan Road, Guangzhou, 510080, China

    Yan-ping Han, Zheng-yu Li, Bao-chuan Li, Xi Sun, Cheng-cheng Zhu, Xiao-ting Ling, Huan-qin Zheng, Zhong-dao Wu & Zhi-yue Lv

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

    Yan-ping Han, Zheng-yu Li, Bao-chuan Li, Xi Sun, Cheng-cheng Zhu, Xiao-ting Ling, Huan-qin Zheng, Zhong-dao Wu & Zhi-yue Lv

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  1. Yan-ping Han
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Correspondence to Zhong-dao Wu or Zhi-yue Lv.

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Han, Yp., Li, Zy., Li, Bc. et al. Molecular cloning and characterization of a cathepsin B from Angiostrongylus cantonensis . Parasitol Res 109, 369–378 (2011). https://doi.org/10.1007/s00436-011-2264-0

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  • DOI: https://doi.org/10.1007/s00436-011-2264-0

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