Purification and characterization of a hemagglutinin from the red alga, Gracilariopsis chorda, from Japan

Hirotaka Kakita¹, Naoki Yanaoka¹, Hideki Obika²

¹Graduate School of Integrated Basic Sciences, Nihon University, Tokyo, 156-8550, Japan

²Health Research Institute, National Institute of Advanced Industrial Science and Technology, Kagawa, 761-0301, Japan

Hemagglutinins (Protein-based ones are called lectins) are natural bioactive substances, which possess specific carbohydrate binding abilities. Four purified hemagglutinins in Japanese Gracilaria vermiculophylla have already been reported in detail. However, a purified hemagglutinin from Gracilariopsis chorda, which belongs to the same algal family Gracilariaceae as Gracilaria vermiculophylla, has not been reported in detail. Thus, in this study, we carried out purification and characterization of a hemagglutinin in the red alga, Gracilariopsis chorda. Ammonium sulfate concentration and standing time in the salting-out process were optimized to obtain hemagglutinin fractions with high specific activity. The combination first salting-out with 35% saturation for 1 hour and second salting-out with 70% saturation for 18 hour was most effective. A hemagglutinin was purified from an ammonium sulfate precipitation by ion exchange and gel filtration chromatographies. The Gracilariopsis chorda hemagglutinin contained large amounts of hexose and sulfate along with a small amount of protein. It agglutinated rabbit erythrocytes and had no divalent cation requirement for hemagglutination. The hemagglutinating activity was inhibited by complex-type glycoproteins, such as asialofetuin, fetuin and thyroglobulin, but was not inhibited by any of the monosaccharides tested. The results suggested that the Gracilariopsis chorda hemagglutinin was different from terrestrial organisms reported to date. The specific activity of the Gracilariopsis chorda hemagglutinin was higher than that of the high molecular weight Gracilaria vermiculophylla hemagglutin (H-GVH). Thus, and as with Gracilaria vermiculophylla, Gracilariopsis chorda seems to be a suitable source of industrially useful hemagglutinins.