Identification and characterization of serine proteinase inhibitors from Neospora caninum.
Mol Biochem Parasitol
; 136(1): 101-7, 2004 Jul.
Article
em En
| MEDLINE
| ID: mdl-15138071
Two cDNA clones obtained from the Neospora caninum Expressed Sequence Tag project were selected by their homology with the Toxoplasma gondii serine proteinase inhibitor (serpin) gene, TgPI-1 and TgPI-2. One of them, named NcPI-H, showed several premature stop codons. The other cDNA, named NcPI-S, encoded a 79 amino acid protein containing a putative signal peptide and only one non-classical Kazal domain. Two other N. caninum EST sequences (NcEST1 and NcEST2) and one from Eimeria tenella (EtPI-S) were retrieved from the database. Amino acid sequence analysis suggested that NcEST1 and NcEST2 might be the N. caninum counterparts of TgPI-1 and TgPI-2, respectively. EtEST-S, as NcPI-S, is a single domain serpin. The open reading frame encoding the mature version of NcPI-S was expressed as recombinant protein, fused to a 6 histidine tag in Escherichia coli. Specific rabbit antiserum generated against the recombinant NcPI-S was used in immunoblot assays. Bands of 20, 30, 40, and 66-kDa were detected by SDS-PAGE of whole parasite homogenate. In addition, when an anti-TgPI-1 serum was used, bands of 25 and 35-kDa were detected indicating that there is no cross-reactivity between both serpins, and showing as well, the presence of another putative serpin in N. caninum. The recombinant protein NcPI-S, inhibited bacterial subtilisin completely, and showed lower inhibitory capacity on human neutrophil elastase, animal trypsin, and chymotrypsin, suggesting differences in effectiveness.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Inibidores de Serina Proteinase
/
Neospora
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
Idioma:
En
Revista:
Mol Biochem Parasitol
Ano de publicação:
2004
Tipo de documento:
Article
País de afiliação:
Argentina
País de publicação:
Holanda