Identification of active Plasmodium falciparum calpain to establish screening system for Pf-calpain-based drug development

Byoung Yul Soh, Hyun Ok Song, Yoonji Lee, Junghyun Lee, Kusuma Kaewintajuk, Binna Lee, Yun Young Choi, Jeong Hoon Cho, Sun Choi, Hyun Park

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15 Scopus citations


Background: With the increasing resistance of malaria parasites to available drugs, there is an urgent demand to develop new anti-malarial drugs. Calpain inhibitor, ALLN, is proposed to inhibit parasite proliferation by suppressing haemoglobin degradation. This provides Plasmodium calpain as a potential target for drug development. Pf-calpain, a cysteine protease of Plasmodium falciparum, belongs to calpain-7 family, which is an atypical calpain not harboring Ca§ssup§2+§esup§-binding regulatory motifs. In this present study, in order to establish the screening system for Pf-calpain specific inhibitors, the active form of Pf-calpain was first identified. Methods. Recombinant Pf-calpain including catalytic subdomain IIa (rPfcal-IIa) was heterologously expressed and purified. Enzymatic activity was determined by both fluorogenic substrate assay and gelatin zymography. Molecular homology modeling was carried out to address the activation mode of Pf-calpain in the aspect of structural moiety. Results: Based on the measurement of enzymatic activity and protease inhibitor assay, it was found that the active form of Pf-calpain only contains the catalytic subdomain IIa, suggesting that Pf-calpain may function as a monomeric form. The sequence prediction indicates that the catalytic subdomain IIa contains all amino acid residues necessary for catalytic triad (Cys-His-Asn) formation. Molecular modeling suggests that the Pf-calpain subdomain IIa makes an active site, holding the catalytic triad residues in their appropriate orientation for catalysis. The mutation analysis further supports that those amino acid residues are functional and have enzymatic activity. Conclusion: The identified active form of Pf-calpain could be utilized to establish high-throughput screening system for Pf-calpain inhibitors. Due to its unique monomeric structural property, Pf-calpain could be served as a novel anti-malarial drug target, which has a high specificity for malaria parasite. In addition, the monomeric form of enzyme may contribute to relatively simple synthesis of selective inhibitors.

Original languageEnglish
Article number47
JournalMalaria Journal
Issue number1
StatePublished - 2013

Bibliographical note

Funding Information:
This work was supported by the National Research Foundation of Korea (NRF) grants funded by the Korea government (MEST) (No. 2012–0005624 and No. 2011–0028885).


  • Anti-malarial drug
  • Calpain
  • Cysteine protease
  • Malaria
  • Monomer
  • Plasmodium falciparum
  • Protease inhibitor


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