title: Ascaris lumbricoides β carbonic anhydrase: a potential target enzyme for treatment of ascariasis.
abstract: 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 kcat of 6.0 × 10(5) s(-1) and kcat/KM 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.
authors.affiliation: Department of Anatomy, School of Medicine, University of Tampere, Tampere, Finland. zolfaghari.emameh.reza.x@student.uta.fi.|BioMediTech, University of Tampere, Tampere, Finland. zolfaghari.emameh.reza.x@student.uta.fi.|Fimlab Laboratories Ltd and Tampere University Hospital, Tampere, Finland. zolfaghari.emameh.reza.x@student.uta.fi.
authors.forename: Reza
authors.initials: R
authors.lastname: Zolfaghari Emameh
authors.affiliation: Department of Anatomy, School of Medicine, University of Tampere, Tampere, Finland. marianne.kuuslahti@staff.uta.fi.
authors.forename: Marianne
authors.initials: M
authors.lastname: Kuuslahti
authors.affiliation: Dipartimento di Chimica, Laboratorio di Chimica Bioinorganica, Universita' degli Studi di Firenze, Sesto Fiorentino, Firenze, Italy. daniela.vullo@unifi.it.|Neurofarba Department, Sezione di Scienze Farmaceutiche e Nutraceutiche, Universita' degli Studi di Firenze, Sesto Fiorentino, Firenze, Italy. daniela.vullo@unifi.it.
authors.forename: Daniela
authors.initials: D
authors.lastname: Vullo
authors.affiliation: Department of Anatomy, School of Medicine, University of Tampere, Tampere, Finland. Barker.Harlan.R@student.uta.fi.
authors.forename: Harlan R
authors.initials: HR
authors.lastname: Barker
authors.affiliation: Dipartimento di Chimica, Laboratorio di Chimica Bioinorganica, Universita' degli Studi di Firenze, Sesto Fiorentino, Firenze, Italy. claudiu.supuran@unifi.it.|Neurofarba Department, Sezione di Scienze Farmaceutiche e Nutraceutiche, Universita' degli Studi di Firenze, Sesto Fiorentino, Firenze, Italy. claudiu.supuran@unifi.it.
authors.forename: Claudiu T
authors.initials: CT
authors.lastname: Supuran
authors.affiliation: Department of Anatomy, School of Medicine, University of Tampere, Tampere, Finland. seppo.parkkila@staff.uta.fi.|Fimlab Laboratories Ltd and Tampere University Hospital, Tampere, Finland. seppo.parkkila@staff.uta.fi.
authors.forename: Seppo
authors.initials: S
authors.lastname: Parkkila
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001196:Ascariasis / Q000469:parasitology*; D017164:Ascaris lumbricoides / Q000201:enzymology*; D002256:Carbonic Anhydrases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D016415:Sequence Alignment
chemical_list: D002256:Carbonic Anhydrases