A novel signal sequence negative multimeric glycosomal protein required for cell cycle progression of Leishmania donovani parasites

Publication date: August 2018Source: Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, Volume 1865, Issue 8Author(s): Kavita Ahuja, Mirza Adil Beg, Ruby Sharma, Ajay Saxena, Nilofer Naqvi, Niti Puri, Pradeep Kumar Rai, Ashok Chaudhury, Robert Duncan, Poonam Salotra, Hira Nakhasi, Angamuthu SelvapandiyanAbstractExpression of the intracellular form amastigote specific genes in the Leishmania donovani parasite plays a major role in parasite replication in the macrophage. In the current work, we have characterized a novel hypothetical gene, Ld30b that is specifically transcribed in the intracellular stage of the parasite. The recombinant Ld30b protein exists as a pentamer in solution as identified by native-PAGE and size exclusion gel chromatography. Structural analysis using circular dichroism and molecular modeling indicate that Ld30b belongs to family of cAMP-dependent protein kinase type I-alpha regulatory subunit. Co-localization immunofluorescence microscopy and western blot analyses (using anti-Ld30b antibody and anti-hypoxanthine-guanine phosphoribosyl transferase, a glycosome marker) on the isolated parasite glycosome organelle fractions show that Ld30b is localized in glycosome, though lacked a glycosome targeting PTS1/2 signal in the protein sequence. Episomal expression of Ld30b in the parasite caused the arrest of promastigotes and amastigotes growth in vitro. Cell cycle analysis using flow cytometry indicates that these parasites are arrested in ‘sub ...
Source: Biochimica et Biophysica Acta (BBA) Molecular Cell Research - Category: Molecular Biology Source Type: research