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Molecular and Functional Analyses of a Novel Secretory Nuclease from the Human Pathogen Leishmania donovani
Author(s) -
Joshi Manju B,
Owings Joshua P,
Baer J Austin,
Dwyer Dennis M
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.5.a277
Subject(s) - biology , amastigote , nuclease , leishmania , leishmania donovani , secretory protein , gene , microbiology and biotechnology , biochemistry , visceral leishmaniasis , parasite hosting , leishmaniasis , genetics , world wide web , computer science
Leishmania are protozoan pathogens of humans that reside/multiply as promastigotes in the gut of their sandfly vectors and as amastigotes in the phago‐lysosomal compartments of infected macrophages. These parasites are purine auxotrophs and thus must salvage such essential nutrients from their hosts. Here we report the identification, characterization and homologous episomal‐expression of a gene ( LdNUC ) encoding a unique 35‐kDa Class‐I “secretory nuclease” in L. donovani . Structural features of the LdNUC deduced protein are consistent with those of secretory proteins. LdNUC mRNA and secretory nuclease activity were constitutively expressed by both promastigotes and amastigotes forms of this parasite. In Western blots and immunoprecipitation (IP) assays, an anti‐ Ld NUC antibody recognized both the native 35 kDa secretory enzyme and the Ld NUC expressed enzyme. Results of coupled IP‐enzyme activity analyses demonstrated that this secretory enzyme could hydrolyze a variety of synthetic polynucleotides as well as, both ss‐ and ds‐DNAs and RNA. Results of PCR‐, Southern‐, Western‐ and zymogram‐analyses all indicated that homologs of LdNUC are conserved amongst all pathogenic species of Leishmania . These observations suggest that this “secretory” nuclease could function away from the parasite, to hydrolyze host‐derived nucleic acids to satisfy their essential purine requirements. Thus, the Ld NUC might play critical role(s) in facilitating the survival, growth and development of these important human pathogens. This study was supported by the Intramural Research Program of the DIR, NIAID, NIH.