Ascaris lumbricoides β carbonic anhydrase: a potential target enzyme for treatment of ascariasis
Zolfaghari, Emameh Reza; Kuuslahti, Marianne; Vullo, Daniela; Barker, Harlan; Supuran, Claudiu; Parkkila, Seppo (2015)
2015
Parasites & Vectors 8 141
479
BioMediTech - BioMediTech
Lääketieteen yksikkö - School of Medicine
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Julkaisun pysyvä osoite on
https://urn.fi/URN:NBN:fi:uta-201510022329
https://urn.fi/URN:NBN:fi:uta-201510022329
Kuvaus
BioMed Central open access
Tiivistelmä
Background
A parasitic roundworm, Ascaris lumbricoides, is the causative agent of ascariasis, with approximately 760 million cases around the world. Helminthic infections occur with a high prevalence mostly in tropical and developing xcountries. Therefore, design of affordable broad-spectrum anti-helminthic agents against a variety of pathogens, including not only A. lumbricoides but also hookworms and whipworms, is desirable. Beta carbonic anhydrases (β-CAs) are considered promising targets of novel anthelminthics because these enzymes are present in various parasites, while completely absent in vertebrates.
Methods
In this study, we identified an A. lumbricoides β-CA (AIBCA) protein from protein sequence data using bioinformatics tools. We used computational biology resources and methods (including InterPro, CATH/Gene3D, KEGG, and METACYC) to analyze AlBCA and define potential roles of this enzyme in biological pathways. The AlBCA gene was cloned into pFastBac1, and recombinant AIBCA was produced in sf-9 insect cells. Kinetics of AlBCA were analyzed by a stopped-flow method.
Results
Multiple sequence alignment revealed that AIBCA contains the two sequence motifs, CXDXR and HXXC, typical for β-CAs. Recombinant AIBCA showed significant CA catalytic activity with k cat of 6.0 × 10 5 s −1 and k cat /K M of 4.3 × 10 7 M −1 s −1 . The classical CA inhibitor, acetazolamide, showed an inhibition constant of 84.1 nM. Computational modeling suggests that the molecular architecture of AIBCA is highly similar to several other known β-CA structures. Functional predictions suggest that AIBCA might play a role in bicarbonate-mediated metabolic pathways, such as gluconeogenesis and removal of metabolically produced cyanate.
Conclusions
These results open new avenues to further investigate the precise functions of β-CAs in parasites and suggest that novel β-CA specific inhibitors should be developed and tested against helminthic diseases.
A parasitic roundworm, Ascaris lumbricoides, is the causative agent of ascariasis, with approximately 760 million cases around the world. Helminthic infections occur with a high prevalence mostly in tropical and developing xcountries. Therefore, design of affordable broad-spectrum anti-helminthic agents against a variety of pathogens, including not only A. lumbricoides but also hookworms and whipworms, is desirable. Beta carbonic anhydrases (β-CAs) are considered promising targets of novel anthelminthics because these enzymes are present in various parasites, while completely absent in vertebrates.
Methods
In this study, we identified an A. lumbricoides β-CA (AIBCA) protein from protein sequence data using bioinformatics tools. We used computational biology resources and methods (including InterPro, CATH/Gene3D, KEGG, and METACYC) to analyze AlBCA and define potential roles of this enzyme in biological pathways. The AlBCA gene was cloned into pFastBac1, and recombinant AIBCA was produced in sf-9 insect cells. Kinetics of AlBCA were analyzed by a stopped-flow method.
Results
Multiple sequence alignment revealed that AIBCA contains the two sequence motifs, CXDXR and HXXC, typical for β-CAs. Recombinant AIBCA showed significant CA catalytic activity with k cat of 6.0 × 10 5 s −1 and k cat /K M of 4.3 × 10 7 M −1 s −1 . The classical CA inhibitor, acetazolamide, showed an inhibition constant of 84.1 nM. Computational modeling suggests that the molecular architecture of AIBCA is highly similar to several other known β-CA structures. Functional predictions suggest that AIBCA might play a role in bicarbonate-mediated metabolic pathways, such as gluconeogenesis and removal of metabolically produced cyanate.
Conclusions
These results open new avenues to further investigate the precise functions of β-CAs in parasites and suggest that novel β-CA specific inhibitors should be developed and tested against helminthic diseases.
Kokoelmat
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