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Cross-talk between malarial cysteine proteases and falstatin: the BC loop as a hot-spot target.

Sundararaj S, Saxena AK, Sharma R, Vashisht K, Sharma S, Anvikar A, Dixit R, Rosenthal PJ, Pandey KC - PLoS ONE (2014)

Bottom Line: Using site-directed mutagenesis, hemoglobin hydrolysis assays and peptide inhibition studies, we demonstrate that the BC loop, but not the DE or FG loops, inhibits cysteine proteases of P. falciparum and P. vivax via hydrogen bonds.These results suggest that the BC loop of falstatin acts as a hot-spot target for inhibiting malarial cysteine proteases.This finding suggests new strategies for the development of anti-malarial agents based on protease-inhibitor interactions.

View Article: PubMed Central - PubMed

Affiliation: Host-Parasite Interaction Biology Group, National Institute of Malaria Research, Indian Council of Medical Research, Dwarka, New Delhi, India.

ABSTRACT
Cysteine proteases play a crucial role in the development of the human malaria parasites Plasmodium falciparum and Plasmodium vivax. Our earlier studies demonstrated that these enzymes are equipped with specific domains for defined functions and further suggested the mechanism of activation of cysteine proteases. The activities of these proteases are regulated by a new class of endogenous inhibitors of cysteine proteases (ICPs). Structural studies of the ICPs of Trypanosoma cruzi (chagasin) and Plasmodium berghei (PbICP) indicated that three loops (termed BC, DE, and FG) are crucial for binding to target proteases. Falstatin, an ICP of P. falciparum, appears to play a crucial role in invasion of erythrocytes and hepatocytes. However, the mechanism of inhibition of cysteine proteases by falstatin has not been established. Our study suggests that falstatin is the first known ICP to function as a multimeric protein. Using site-directed mutagenesis, hemoglobin hydrolysis assays and peptide inhibition studies, we demonstrate that the BC loop, but not the DE or FG loops, inhibits cysteine proteases of P. falciparum and P. vivax via hydrogen bonds. These results suggest that the BC loop of falstatin acts as a hot-spot target for inhibiting malarial cysteine proteases. This finding suggests new strategies for the development of anti-malarial agents based on protease-inhibitor interactions.

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Alignment of falstatin with other ICPs.A multiple sequence alignment was performed with falstatin and ICPs from Cryptosporidium parvum (cryptostatin), Trypanosoma cruzi (chagasin), and Plasmodium berghei (PbICP). This alignment predicted four major loop regions; L0, BC (L2), DE (L4) and FG (L6) in falstatin. The peptides used in this study are underlines, and residues mutated in the described studies are in bold type.
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pone-0093008-g001: Alignment of falstatin with other ICPs.A multiple sequence alignment was performed with falstatin and ICPs from Cryptosporidium parvum (cryptostatin), Trypanosoma cruzi (chagasin), and Plasmodium berghei (PbICP). This alignment predicted four major loop regions; L0, BC (L2), DE (L4) and FG (L6) in falstatin. The peptides used in this study are underlines, and residues mutated in the described studies are in bold type.

Mentions: Structural studies of chagasin and PbICP led to the identification of crucial loops in the ICP family of proteins. Falstatin and its plasmodial homologues have molecular masses much higher than those of other ICPs, and so it was unclear if the structural determinants of inhibition by falstatin are similar to those of non plasmodial ICPs [11], [12]. Our first objective was to identify and characterize the determinant of inhibition in falstatin. We aligned the sequences of falstatin and PbICP with those of other ICPs. Falstatin and it plasmodial homologues have long stretches of amino acids that are absent in chagasin and homologues from Leishmania mexicana and Cryptosporidium parvum[11], [12], [32] (Fig. 1). Overall, the sequence similarity between falstatin and chagasin is ∼20%, and that between falstatin and PbICP is ∼38%. Based on homology with the solved structures of chagasin and PbICP [12], [15], we identified four major loops, labeled as L0, BC (L2), DE (L4) and FG (L6) (Fig. 1).


Cross-talk between malarial cysteine proteases and falstatin: the BC loop as a hot-spot target.

Sundararaj S, Saxena AK, Sharma R, Vashisht K, Sharma S, Anvikar A, Dixit R, Rosenthal PJ, Pandey KC - PLoS ONE (2014)

Alignment of falstatin with other ICPs.A multiple sequence alignment was performed with falstatin and ICPs from Cryptosporidium parvum (cryptostatin), Trypanosoma cruzi (chagasin), and Plasmodium berghei (PbICP). This alignment predicted four major loop regions; L0, BC (L2), DE (L4) and FG (L6) in falstatin. The peptides used in this study are underlines, and residues mutated in the described studies are in bold type.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3974720&req=5

pone-0093008-g001: Alignment of falstatin with other ICPs.A multiple sequence alignment was performed with falstatin and ICPs from Cryptosporidium parvum (cryptostatin), Trypanosoma cruzi (chagasin), and Plasmodium berghei (PbICP). This alignment predicted four major loop regions; L0, BC (L2), DE (L4) and FG (L6) in falstatin. The peptides used in this study are underlines, and residues mutated in the described studies are in bold type.
Mentions: Structural studies of chagasin and PbICP led to the identification of crucial loops in the ICP family of proteins. Falstatin and its plasmodial homologues have molecular masses much higher than those of other ICPs, and so it was unclear if the structural determinants of inhibition by falstatin are similar to those of non plasmodial ICPs [11], [12]. Our first objective was to identify and characterize the determinant of inhibition in falstatin. We aligned the sequences of falstatin and PbICP with those of other ICPs. Falstatin and it plasmodial homologues have long stretches of amino acids that are absent in chagasin and homologues from Leishmania mexicana and Cryptosporidium parvum[11], [12], [32] (Fig. 1). Overall, the sequence similarity between falstatin and chagasin is ∼20%, and that between falstatin and PbICP is ∼38%. Based on homology with the solved structures of chagasin and PbICP [12], [15], we identified four major loops, labeled as L0, BC (L2), DE (L4) and FG (L6) (Fig. 1).

Bottom Line: Using site-directed mutagenesis, hemoglobin hydrolysis assays and peptide inhibition studies, we demonstrate that the BC loop, but not the DE or FG loops, inhibits cysteine proteases of P. falciparum and P. vivax via hydrogen bonds.These results suggest that the BC loop of falstatin acts as a hot-spot target for inhibiting malarial cysteine proteases.This finding suggests new strategies for the development of anti-malarial agents based on protease-inhibitor interactions.

View Article: PubMed Central - PubMed

Affiliation: Host-Parasite Interaction Biology Group, National Institute of Malaria Research, Indian Council of Medical Research, Dwarka, New Delhi, India.

ABSTRACT
Cysteine proteases play a crucial role in the development of the human malaria parasites Plasmodium falciparum and Plasmodium vivax. Our earlier studies demonstrated that these enzymes are equipped with specific domains for defined functions and further suggested the mechanism of activation of cysteine proteases. The activities of these proteases are regulated by a new class of endogenous inhibitors of cysteine proteases (ICPs). Structural studies of the ICPs of Trypanosoma cruzi (chagasin) and Plasmodium berghei (PbICP) indicated that three loops (termed BC, DE, and FG) are crucial for binding to target proteases. Falstatin, an ICP of P. falciparum, appears to play a crucial role in invasion of erythrocytes and hepatocytes. However, the mechanism of inhibition of cysteine proteases by falstatin has not been established. Our study suggests that falstatin is the first known ICP to function as a multimeric protein. Using site-directed mutagenesis, hemoglobin hydrolysis assays and peptide inhibition studies, we demonstrate that the BC loop, but not the DE or FG loops, inhibits cysteine proteases of P. falciparum and P. vivax via hydrogen bonds. These results suggest that the BC loop of falstatin acts as a hot-spot target for inhibiting malarial cysteine proteases. This finding suggests new strategies for the development of anti-malarial agents based on protease-inhibitor interactions.

Show MeSH
Related in: MedlinePlus