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Characterization and mRNA expression analysis of PI31, an endogenous proteasome inhibitor from Schistosoma mansoni

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Abstract

The proline-rich inhibitor of 31 kDa (PI31) is highly conserved through metazoan evolution, and its activity in the proteasome inhibition is well-established although the precise mechanism of inhibition is unclear. The coding DNA sequence of Schistosoma mansoni PI31 (SmPI31) was cloned, and the recombinant protein was expressed in bacterial system. The correct amino acid sequence was confirmed by mass spectrometry and circular dichroism suggests that SmPI31 contains both α-helix and non-structured regions. Inhibition assays, using the Suc-Leu-Leu-Val-Tyr-4-MCA substrate for proteasome degradation, showed that the S. mansoni proteasome may be regulated by the inhibitory activity of SmPI31. A gene expression assay using qRT-PCR at various stages during the S. mansoni life cycle has shown that SmPI31 transcripts are expressed in all studied stages, suggesting that PI31 plays an important role during the developmental processes of the parasite. In this study first evidence is presented that PI31 has a conserved structure and plays a role as proteasome inhibitor in adult worms and it is expressed through life cycle.

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Abbreviations

Suc-Leu-Leu-Val-Tyr-MCA:

Succinyl-Leucine-Leucine-Valine-Tyrosine-4-methyl-coumaryl-7-amide

UVCD:

Ultra-violet circular dichroism

IPTG:

Isopropyl-β-d-thiogalactopyranoside

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Acknowledgments

The authors thank the Brazilian research agencies: FAPESP (Fundação de Amparo a Pesquisa do Estado de São Paulo) and CAPES (Coordenacao de Amparo ao Pessoal de Nivel Superior). We are also grateful to Elenice A. Macedo and Olinda Mara Brigato for their technical assistance.

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Authors and Affiliations

  1. Department of Biochemistry and Immunology, FMRP-USP, Av. Bandeirantes, 3900, Ribeirão Preto, São Paulo, Brazil, 14040-900

    Carla Botelho-Machado, F. J. Cabral, C. S. Soares, E. B. C. Moreira, E. R. Morais, L. G. Magalhães & V. Rodrigues

  2. Institute of Biological Sciences (ICEB), Federal University of Ouro Preto, Ouro Preto, Minas Gerais, Brazil

    M. S. Gomes & R. Guerra-Sá

  3. Protein Chemistry Center, FH-HCFMRP-USP, Ribeirão Preto, São Paulo, Brazil

    J. C. Rosa

  4. Department of Chemistry, FFCLRP-USP, Av. Bandeirantes, 3900, Ribeirão Preto, São Paulo, Brazil

    R. Ruller & R. J. Ward

Authors
  1. Carla Botelho-Machado
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  2. F. J. Cabral
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  3. C. S. Soares
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  4. E. B. C. Moreira
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  5. E. R. Morais
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  6. L. G. Magalhães
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  7. M. S. Gomes
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  8. R. Guerra-Sá
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  9. J. C. Rosa
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  10. R. Ruller
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  11. R. J. Ward
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  12. V. Rodrigues
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Correspondence to Carla Botelho-Machado.

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Botelho-Machado, C., Cabral, F.J., Soares, C.S. et al. Characterization and mRNA expression analysis of PI31, an endogenous proteasome inhibitor from Schistosoma mansoni . Parasitol Res 107, 1163–1171 (2010). https://doi.org/10.1007/s00436-010-1984-x

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  • DOI: https://doi.org/10.1007/s00436-010-1984-x

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