Heterologous expression, purification and characterization of L-type lectin homologue from Leishmania donovani
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Date
2015
Journal Title
Journal ISSN
Volume Title
Publisher
Elsevier
Abstract
Leishmaniasis, a disease of the developing world affects about 12 million people and has limited therapeutic interventions available. L-type lectins, Endoplasmic Reticulum Golgi Intermediate Compartment/Vesicular Integral Proteins (ERGIC-53/VIP36) are involved in protein sorting in luminal compartments of animal cells and are important for parasite biology. A lectin homologue was identified through a bioinformatics analysis of Leishmania genome and it was found to have N-terminal conserved carbohydrate recognition domain (CRD) and a unique C-terminal region rich in repetitive amino acids and a poly glutamine tract. The N-terminal CRD region was cloned and expressed in Escherichia coli, but gave an insoluble expression which was re-solubilized by on column refolding. The fold integrity was checked through CD, fluorescence and functional assay of hemagglutination activity using rabbit erythrocyte. Bioinformatics analysis identified 15 members from Tritryps (Leishmania spp., Trypanosoma spp.) and they separate out as a distinct clade in the global phylogenetic analysis of all ERGIC-53/VIP36 sequences downloaded from Uniprot. Our analysis shows that the extended C-terminal regions with repeats is unique to Tritryps and this repeat pattern is different in sequences from Leishmania spp. and Trypanosoma spp. and all these features make this protein an interesting candidate for further detailed studies. ? 2015 The Author. Published by Elsevier B.V.
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Keywords
lectin bioinformatics, fluorescence, gene amplification, hemagglutination, Leishmania donovani, mass spectrometry, nonhuman, phylogeny, polyacrylamide gel electrophoresis, polymerase chain reaction, protein analysis, protein expression, protein purification, protein refolding, rabbit, sequence analysis, Trypanosoma
Citation
Singh, V., Nair, D. N., Kaushal, R. S., Kumar, M., Pappachan, A., & Singh, D. D. (2015). Heterologous expression, purification and characterization of L-type lectin homologue from Leishmania donovani. Biotechnology Reports, 8, 81-87. doi: 10.1016/j.btre.2015.09.004