Cloning, expression, purification and characterization of Leishmania tropica PDI-2 protein

Abstract: In Leishmania species, protein disulfide isomerase (PDI) is an essential enzyme that catalyzes thiol-disulfide interchange. The present work describes the isolation, cloning, sequencing and expression of the pdI-2 gene. Initially, the gene was amplified from L. tropica genomic DNA by PCR using specific primers before cloning into the expression vector pET-15b. The construct pET/pdI-2 was transformed into BL21 (DE3) cells and induced for the protein expression. SDS-PAGE and western blot analysis showed that the expressed protein is about 51 kDa. Cloned gene sequence analysis revealed that the deduced amino acid sequence showed significant homology with those of several parasites PDIs. Finally, recombinant protein was purified with a metal-chelating affinity column. The putative protein was confirmed as a thiol - disulfide oxidoreductase by detecting its activity in an oxidoreductase assay. Assay result of assay suggested that the PDI-2 protein is required for both oxidation and reduction of disulfide bonds in vitro. Antibodies reactive with this 51 kDa protein were detected by Western blot analysis in sera from human infected with L. tropica. This work describes for the first time the enzymatic activity of recombinant L. tropica PDI-2 protein and suggests a role for this protein as an antigen for the detection of leishmaniasis infection.

Object Details

• Location: Deutsche Nationalbibliothek Frankfurt am Main
• Extent: Online-Ressource
• Language: Englisch

References and Relationships

• Bibliographic citation: Cloning, expression, purification and characterization of Leishmania tropica PDI-2 protein ; volume:11 ; number:1 ; year:2016 ; pages:166-176 ; extent:11 ; Open life sciences ; 11, Heft 1 (2016), 166-176 (gesamt 11)

Contributors, Places and Time

• Creator: Ali, Dina ; Abbady, Abdul-Qader ; Kweider, Mahmoud ; Soukkarieh, Chadi

Further information

• DOI: 10.1515/biol-2016-0022
• URN: urn:nbn:de:101:1-2409201629367.987874512786
• Rights: Open Access; Der Zugriff auf das Objekt ist unbeschränkt möglich.
• Last update: 15.08.2025, 7:27 AM CEST