Cloning, expression and characterization of novel lectin from Oryza sativa

초록

A lectin gene homolog of Oryza sativa was successfully cloned and expressed in Escherichia coli. The deduced amino acid sequence of the protein product showed a significant similarity with known chitin-binding lectins. Most of the recombinant lectin was found in an insoluble aggregated form as inclusion bodies and only a small part was in the culture medium in a soluble active form. Functional recombinant lectin was recovered from the inclusion bodies by solubilization with 8 M urea in Tris/HCl buffer, pH 7.0 and renaturation by 10-fold dilution in the same buffer. The recombinant lectin with His-tag was simply purified to homogeneity by the process of affinity chromatography and was obtained with a yield of 6-8 mg/L culture. The recombinant lectin was a homo-dimer composed of 22 kDa. The hemagglutination activity of the recombinant lectin was optimal at pH 4.0-7.0 and it was very sensitive to inhibition by N-acetylneuraminic acid and thyroglobulin.

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