Cell Stress and Chaperones

, Volume 18, Issue 2, pp 171–181 | Cite as

Ddi1-like protein from Leishmania major is an active aspartyl proteinase

  • María J. Perteguer
  • Paulino Gómez-Puertas
  • Carmen Cañavate
  • Francehuli Dagger
  • Teresa Gárate
  • Elizabeth Valdivieso
Original Paper

Abstract

Eukaryotic cells respond to DNA damage by activating damage checkpoint pathways, which arrest cell cycle progression and induce gene expression. We isolated a full-length cDNA encoding a 49-kDa protein from Leishmania major, which exhibited significant deduced amino acid sequence homology with the annotated Leishmania sp. DNA damage-inducible (Ddi1-like) protein, as well as with the Ddi1 protein from Saccharomyces cerevisiae. In contrast to the previously described Ddi1 protein, the protein from L. major displays three domains: (1) an NH2-terminal ubiquitin like; (2) a COOH terminal ubiquitin-associated; (3) a retroviral aspartyl proteinase, containing the typical D[S/T]G signature. The function of the L. major Ddi1-like recombinant protein was investigated after expression in baculovirus/insect cells and biochemical analysis, revealing preferential substrate selectivity for aspartyl proteinase A2 family substrates, with optimal activity in acidic conditions. The proteolytic activity was inhibited by aspartyl proteinase inhibitors. Molecular modeling of the retroviral domain of the Ddi1-like Leishmania protein revealed a dimer structure that contained a double Asp-Ser-Gly-Ala amino acid sequence motif, in an almost identical geometry to the exhibited by the homologous retroviral aspartyl protease domain of yeast Ddi1 protein. Our results indicate that the isolated Ddi1-like protein is a functional aspartyl proteinase in L. major, opening possibility to be considered as a potential target for novel antiparasitic drugs.

Keywords

Leishmania Aspartyl proteinase Ddi1 Ubiquitin receptor proteins Retroviral protease Cloning 

Notes

Acknowledgments

This study was supported by the AECI grant no. A/023788/09 to TG and EV, and the Spanish Ministry of Science and Innovation and the Instituto de Salud Carlos III within the Network of Tropical Diseases Research (RICET RD06/0021/0019 and RD06/0021/0009). We thank Biomol-Informatics SL (www.biomol-informatics.com) for bioinformatics consulting.

Supplementary material

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Copyright information

© Cell Stress Society International 2012

Authors and Affiliations

  • María J. Perteguer
    • 1
  • Paulino Gómez-Puertas
    • 2
  • Carmen Cañavate
    • 3
  • Francehuli Dagger
    • 4
  • Teresa Gárate
    • 1
  • Elizabeth Valdivieso
    • 4
    • 5
  1. 1.Servicio de Parasitología, Centro Nacional de MicrobiologíaInstituto de Salud Carlos IIIMadridSpain
  2. 2.Centro de Biología Molecular “Severo Ochoa” (CSIC-UAM) Campus UAMMadridSpain
  3. 3.World Health Organization Collaborating Centre for Leishmaniasis, Servicio de Parasitología, Centro Nacional de MicrobiologíaInstituto de Salud Carlos IIIMadridSpain
  4. 4.Laboratorio de Biología Celular de Parásitos, Instituto de Biología ExperimentalUniversidad Central de VenezuelaCaracasVenezuela
  5. 5.Instituto de Biología ExperimentalCaracasVenezuela

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