Expression in bacteria of the gene encoding the gp43 antigen of paracoccidioides brasiliensis: immunological reactivity of the recombinant fusion proteins.
gp43 is the major diagnostic antigen of Paracoccidioides brasiliensis, the agent of paracoccidioidomycosis (PCM) in humans. In the present study, cDNA of the gp43 gene (PbGP43) was obtained by reverse transcriptase PCR, inserted into a pGEX vector in frame with the glutathione S-transferase (GST) gene, and expressed in Escherichia coli as inclusion bodies. Immunoblotting showed that all sera from patients with chronic pulmonary and acute lymphatic forms of PCM reacted with the recombinant fusion protein of the mature gp43 (381 amino acids). Reactivity with fusion proteins containing subfragments of the N-terminal, internal, or C-terminal regions occurred eventually, and the C-terminal region was the most antigenic. Lack of reactivity with the subfragments may be due to the conformational nature of the gp43 epitopes. Sera from patients with aspergillosis, candidiasis, and histoplasmosis did not react with the gp43-GST fusion protein. Our results suggest that recombinant gp43 corresponding to the processed antigen can be a useful tool in the diagnosis of PCM.
Diniz
Susana N
SN
Departamento de Microbiologia, Imunologia e Parasitologia da Universidade Federal de São Paulo, São Paulo, SP 04023-062, Brazil.
Carvalho
Kátia C
KC
Cisalpino
Patrícia S
PS
Silveira
José F
JF
Travassos
Luiz R
LR
Puccia
Rosana
R
D000818:Animals; D000946:Antigens, Fungal / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D004926:Escherichia coli / Q000235:genetics; D015151:Immunoblotting; D010228:Paracoccidioides / Q000276:immunology*; D010229:Paracoccidioidomycosis / Q000175:diagnosis*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000276:immunology*
D000946:Antigens, Fungal; D011993:Recombinant Fusion Proteins