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Clinical and Diagnostic Laboratory Immunology, November 2002, p. 1200-1204, Vol. 9, No. 6
1071-412X/02/$04.00+0     DOI: 10.1128/CDLI.9.6.1200-1204.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Expression in Bacteria of the Gene Encoding the gp43 Antigen of Paracoccidioides brasiliensis: Immunological Reactivity of the Recombinant Fusion Proteins

Susana N. Diniz,{dagger} Kátia C. Carvalho, Patrícia S. Cisalpino,{ddagger} José F. Silveira, Luiz R. Travassos, and Rosana Puccia*

Departamento de Microbiologia, Imunologia e Parasitologia da Universidade Federal de São Paulo, São Paulo, SP 04023-062, Brazil

Received 18 April 2002/ Returned for modification 13 June 2002/ Accepted 22 July 2002

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.

* Corresponding author. Mailing address: Disciplina de Biologia Celular, UNIFESP, Rua Botucatu, 862, oitavo andar, São Paulo, SP 04023-062, Brazil. Phone: 55-11-5084-2991. Fax: 55-11-5571-5877. E-mail: rosana{at}ecb.epm.br.

{dagger} Present address: Instituto Ludwig de Pesquisa do CÂncer, São Paulo, Brazil.

{ddagger} Present address: Departamento de Microbiologia do Instituto de Ciências Biológicas Da Universidade Federal de Minas Gerais, Belo Horizonte, MG 31270-901, Brazil.

Clinical and Diagnostic Laboratory Immunology, November 2002, p. 1200-1204, Vol. 9, No. 6
1071-412X/02/$04.00+0     DOI: 10.1128/CDLI.9.6.1200-1204.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.





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