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Expression, purification and characterization of a galectin in the silkworm( Bombyx mori)
Author(s): 
Pages: 806-814
Year: Issue:  7
Journal: Acta Entomologica Sinica

Keyword:  Bombyx morimicrobial challengegalectinrecombinant proteinpolysaccharidesaccharide binding;
Abstract: 【Aim】 To identify a novel galectin gene from the silkworm,Bombyx mori,analyze its sequence and structural characteristics,detect its expression after microbial challenge and measure the binding properties of the expressed recombinant protein with microbe and microbial surface polysaccharides. 【Methods】We used TBlastN to identify a novel galectin gene( named BmGalectin-4)by searching the silkworm genome database,and analyzed its sequence and structural characteristics with bioinformatics tools. The expression of BmGalectin-4 in different tissues in larval silkworm after microbial challenge( inoculation of Pseudomonas aeruginosa,Staphylococcus aureus and Candidiasis albicans,respectively) was detected by RT-PCR. The recombinant protein was produced in Escherichia coli and purified by affinity chromatography. The binding property of BmGalectin-4 with bacteria and fungus was analyzed by ELISA. Western blot was used to detect its binding to polysaccharides. 【Results】Sequence analysis showed that BmGalectin-4 is a tandem repeat galectin,containing two carbohydrate recognition domains. BmGalectin-4 was expressed in fat body,hemocytes and midgut tissues,and its expression level changed significantly after bacterial and fungal challenge. Purified recombinant BmGalectin-4 showed significant binding capacity to Gram-positive and Gram-negative bacteria and fungus. Its binding to microbial surface polysaccharides was highly specific. 【Conclusion】BmGalectin-4 is a typical tandem repeat galectin and might be involved in the silkworm immune responses against microbial pathogens.This study provides the foundation for further functional study of the silkworm galectin.
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