Tesis
Caracterização e avaliação da capacidade imunogênica da enolase de Conidiobolus lamprauges
Fecha
2016-07-29Registro en:
OLIVEIRA, Juçara Tinasi de. Caracterização e avaliação da capacidade imunogênica da enolase de Conidiobolus lamprauges. 2016. 44 f. Tese (Doutorado em Ciências Veterinárias) - Universidade Federal de Mato Grosso, Faculdade de Agronomia, Medicina Veterinária e Zootecnia, Cuiabá, 2016.
Autor
Dutra, Valéria
Nakazato, Luciano
http://lattes.cnpq.br/3898850578198054
http://lattes.cnpq.br/4478191386305454
Dutra, Valéria
501.674.720-20
http://lattes.cnpq.br/4478191386305454
Pescador, Caroline Argenta
958.659.180-87
http://lattes.cnpq.br/5754349416478829
501.674.720-20
638.389.071-91
Pacheco, Richard de Campos
791.476.071-49
http://lattes.cnpq.br/5213594247690553
Silva, Maria Cristina da
866.401.861-87
http://lattes.cnpq.br/3805840098386826
Botton, Sônia de Avila
672.074.720-72
http://lattes.cnpq.br/0814772095155945
Institución
Resumen
Enolase, an important glycolytic enzyme with increased expression at 37°C in
Conidiobolus (C.) lamprauges, performs different functions in other organisms,
among them, adhesion and invasion of hosts. Thus, the identification of enolase as
an immunoreactive protein may contribute to the understanding of these diseases;
provide the standardization of serological tests and also develop vaccines, thus
bringing more effective treatments and reducing the death of animals. The objectives
of this study were to characterize the enolase gene (eno) C. lamprauges and
evaluate its immunogenic capacity towards naturally infected sheep. Eno gene
amplification isolated from C. lamprauges by the oligonucleotides synthesized based
on the sequence of the gene C. lamprauges resulted in a 1305pb fragment. The
deduced aminoacid sequence comprised 434 residues with a predicted molecular
weight of 47,2KDa. In the sequence analysis, we identified the signing of the enolase
gene (LLLKVNQIGTVSES) corresponding to positions 342-345. In the analysis "in
silico" probable regions were detected for epitopes to T and B lymphocytes. The
sequence was cloned into the plasmid pFN6K (HQ) Flexi® Vector (Promega®) and
the protein expressed in Escherichia coli BL21 (DE3) pLysS cells with IPTG
induction. In SDS-PAGE and Western blot, the recombinant eno was detected with
estimated molecular weight of 47kDa. Recombinant IgG anti-enolase was found is in
all animals with conidiobolomycosis by means of the Western blot technique,
demonstrating immunogenicity. In healthy sheep serum, used as a negative control,
and serum from sheep with pythiosis used to evaluate cross-reactivity, bands were
not detected. Therefore, this study demonstrated that recombinant enolase C.
lamprauges is immunogenic against serum of naturally infected sheep with
conidiobolomycosis. The detection of specific anti-enolase antibodies in animals
shows that the protein is a promising candidate antigen for the development of
effective vaccine against this disease.