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Optimizing Expression of Streptococcus pneumoniae Surface Protein a, PspA: Serocross-Reactivity within Families of Antisera Induced Against Clades 1 and 3

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Abstract

Streptococcus pneumoniae is the agent responsible for infections such as pneumonia, otitis media, and meningitis. Among virulence factors, the Pneumococcal surface protein A (PspA) has been shown to be immunogenic and protective in mice, and is thus a good vaccine candidate. PspA has been classified into 6 clades and 3 families. Initially, pspA fragments, clades 1 and 3, were cloned into the pAE-6His expression vector. Proteins were expressed in Escherichia coli BL21(DE3) and purified by affinity and anion exchange chromatographies, with a yield of 11 mg/l of culture. Due to plasmid instability in E. coli, another construct using pspA1 was obtained based on pET-37b(+), which was shown to be stable in E. coli and increased the yield ∼3-fold. Our results show good conditions for scale-up. Sera from immunized mice recognized PspA in total extracts of S. pneumoniae strains: anti-rPspA1p sera recognized native PspA clades 1 (+++), 2 (++) and 4 (+) and anti-rPspA3p sera recognized PspA clades 1 (+), 2 (+), 3 (+++) and 4 (+). The cross-reactivity pattern obtained confirms the notion that proteins from both families should be included for development of a broad-coverage vaccine; lower-cross reactivity between rPspAs of family 2 indicates that it may be necessary to include 2 proteins from this family.

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Acknowledgments

We thank Dr. Mickie Takagi and Dr. Ivan P. Nascimento for assistance during bacterial cultivation and protein analysis. We also thank Ines A. Maurelli, Lourivaldo I. de Souza, Marisa G. Trevelin, Solange R. Silva, Sebastiana V. Oliveira and Sueli G. Pedroza for technical assistance and Fatima A. M. Oliveira for secretarial assistance. This work received financial support from FAPESP, CNPq, and Fundação Butantan.

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  1. Centro de Biotecnologia, Instituto Butantan, Av. Vital Brasil 1500, 05503-900, Sao Paulo, SP, Brazil

    Marcelo Silva, Joaquin Cabrera-Crespo, Maria E. Sbrogio-Almeida, Eliane N. Miyaji, Paulo L. Ho, Luciana C. C. Leite & Alexandre P. Y. Lopes

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  1. Marcelo Silva
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  2. Joaquin Cabrera-Crespo
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  3. Maria E. Sbrogio-Almeida
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  4. Eliane N. Miyaji
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  5. Paulo L. Ho
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  6. Luciana C. C. Leite
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  7. Alexandre P. Y. Lopes
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Correspondence to Alexandre P. Y. Lopes.

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Silva, M., Cabrera-Crespo, J., Sbrogio-Almeida, M.E. et al. Optimizing Expression of Streptococcus pneumoniae Surface Protein a, PspA: Serocross-Reactivity within Families of Antisera Induced Against Clades 1 and 3. Mol Biotechnol 37, 146–154 (2007). https://doi.org/10.1007/s12033-007-0043-z

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  • DOI: https://doi.org/10.1007/s12033-007-0043-z

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