naa a22 4500 510746403 CHVBK 20180411083047.0 cr unu---uuuuu 180411e20130301xx s 000 0 eng 10.1007/s12192-012-0368-9 doi (NATIONALLICENCE)springer-10.1007/s12192-012-0368-9 Ddi1-like protein from Leishmania major is an active aspartyl proteinase [Elektronische Daten] [María Perteguer, Paulino Gómez-Puertas, Carmen Cañavate, Francehuli Dagger, Teresa Gárate, Elizabeth Valdivieso] 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. Cell Stress Society International, 2012 Leishmania nationallicence Aspartyl proteinase nationallicence Ddi1 nationallicence Ubiquitin receptor proteins nationallicence Retroviral protease nationallicence Cloning nationallicence Perteguer María Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220, Madrid, Spain aut Gómez-Puertas Paulino Centro de Biología Molecular "Severo Ochoa” (CSIC-UAM) Campus UAM, Cantoblanco, 28049, Madrid, Spain aut Cañavate Carmen World Health Organization Collaborating Centre for Leishmaniasis, Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220, Madrid, Spain aut Dagger Francehuli Laboratorio de Biología Celular de Parásitos, Instituto de Biología Experimental, Universidad Central de Venezuela, 1041-A, Caracas, Venezuela aut Gárate Teresa Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220, Madrid, Spain aut Valdivieso Elizabeth Laboratorio de Biología Celular de Parásitos, Instituto de Biología Experimental, Universidad Central de Venezuela, 1041-A, Caracas, Venezuela aut Cell Stress and Chaperones Springer Netherlands 18/2(2013-03-01), 171-181 1355-8145 18:2<171 2013 18 12192 https://doi.org/10.1007/s12192-012-0368-9 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s12192-012-0368-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Perteguer María Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220, Madrid, Spain aut NATIONALLICENCE 700 1- Gómez-Puertas Paulino Centro de Biología Molecular "Severo Ochoa” (CSIC-UAM) Campus UAM, Cantoblanco, 28049, Madrid, Spain aut NATIONALLICENCE 700 1- Cañavate Carmen World Health Organization Collaborating Centre for Leishmaniasis, Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220, Madrid, Spain aut NATIONALLICENCE 700 1- Dagger Francehuli Laboratorio de Biología Celular de Parásitos, Instituto de Biología Experimental, Universidad Central de Venezuela, 1041-A, Caracas, Venezuela aut NATIONALLICENCE 700 1- Gárate Teresa Servicio de Parasitología, Centro Nacional de Microbiología, Instituto de Salud Carlos III, Majadahonda, 28220, Madrid, Spain aut NATIONALLICENCE 700 1- Valdivieso Elizabeth Laboratorio de Biología Celular de Parásitos, Instituto de Biología Experimental, Universidad Central de Venezuela, 1041-A, Caracas, Venezuela aut NATIONALLICENCE 773 0- Cell Stress and Chaperones Springer Netherlands 18/2(2013-03-01), 171-181 1355-8145 18:2<171 2013 18 12192 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer