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Cryptopain-1, a cysteine protease of Cryptosporidium parvum, does not require the pro-domain for folding

Published online by Cambridge University Press:  18 December 2008

B.-K. NA ,
J.-M. KANG ,
H.-I. CHEUN ,
S.-H. CHO ,
S.-U. MOON ,
T.-S. KIM  and
W.-M. SOHN
B.-K. NA*
Affiliation:
Department of Parasitology and Institute of Health Sciences, College of Medicine, Gyeongsang National University, Jinju660-751, Korea
J.-M. KANG
Affiliation:
Department of Parasitology and Institute of Health Sciences, College of Medicine, Gyeongsang National University, Jinju660-751, Korea
H.-I. CHEUN
Affiliation:
Department of Malaria and Parasitic Diseases, National Institute of Health, Centers for Disease Control and Prevention, Seoul122-701, Korea
S.-H. CHO
Affiliation:
Department of Malaria and Parasitic Diseases, National Institute of Health, Centers for Disease Control and Prevention, Seoul122-701, Korea
S.-U. MOON
Affiliation:
Department of Malaria and Parasitic Diseases, National Institute of Health, Centers for Disease Control and Prevention, Seoul122-701, Korea
T.-S. KIM
Affiliation:
Department of Parasitology and Inha Research Institute for Medical Sciences, Inha University College of Medicine, Incheon400-751, Korea
W.-M. SOHN
Affiliation:
Department of Parasitology and Institute of Health Sciences, College of Medicine, Gyeongsang National University, Jinju660-751, Korea
*
*Corresponding author: Department of Parasitology and Institute of Health Sciences,College of Medicine, Gyeongsang National University, Jinju660-751, Korea. Tel: +82 55 751 8822. Fax: +82 55 759 4022. E-mail: [email protected]
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Summary

Cryptosporidium parvum is an intracellular protozoan parasite that causes cryptosporidiosis in mammals including humans. In the current study, the gene encoding the cysteine protease of C. parvum (cryptopain-1) was identified and the biochemical properties of the recombinant enzyme were characterized. Cryptopain-1 shared common structural properties with cathepsin L-like papain family enzymes, but lacked a typical signal peptide sequence and contained a possible transmembrane domain near the amino terminus and a unique insert in the front of the mature domain. The recombinant cryptopain-1 expressed in Escherichia coli and refolded to the active form showed typical biochemical properties of cathepsin L-like enzymes. The folding determinant of cryptopain-1 was characterized through multiple constructs with or without different lengths of the pro-domain of the enzyme expressed in E. coli and assessment of their refolding abilities. All constructs, except one that did not contain the full-length mature domain, successfully refolded into the active enzymes, suggesting that cryptopain-1 did not require the pro-domain for folding. Western blot analysis showed that cryptopain-1 was expressed in the sporozoites and the enzyme preferentially degraded proteins, including collagen and fibronectin, but not globular proteins. This suggested a probable role for cryptopain-1 in host cell invasion and/or egression by the parasite.

Keywords

Cryptosporidium parvumcysteine proteasecryptopain-1pro-domainfolding
Type
Research Article
Information
Parasitology , Volume 136 , Issue 2 , February 2009 , pp. 149 - 157
Copyright
Copyright © 2008 Cambridge University Press

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