CLONING, EXPRESSION AND CHARACTERIZATION OF NOVEL LECTIN FROM ORYZA SATIVA |
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Authors: | KWANG-HOON KONG SUNG-GUAN HONG SUN-YOUNG YOO KWANG-SOO LEE HA-HYUNG KIM |
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Affiliation: | Department of Chemistry, College of Natural Science, Chung-Ang University, Dongjak-ku, Seoul 156-756, Korea; College of Pharmacy, Chung-Ang University, Dongjak-ku, Seoul 156-756, Korea |
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Abstract: | 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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