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SFL, a lectin from the marine sponge Stylissa flexibilis was purified by cold ethanol precipitation followed by ion exchange chromatography on DEAE Sepharose column and Sephacryl S-200 gel filtration. SFL is a dimeric glycoprotein of 32 kDa subunits linked by a disulfide bridge with a molecular mass of 64 kDa by SDS-PAGE and 65 kDa by Sephacryl S-200 gel filtration. SFL preferentially agglutinated enzyme treated human A erythrocytes. The activity of lectin was strongly inhibited by monosaccharide d -galactose and glycoproteins asialo-porcine stomach mucin and asialo-fetuin. The lectin was Ca 2 + dependent, stable over a range of pH from 5 to 8, and up to 60 °C for 30 min. The N-terminal amino acid sequence of SFL was also determined and a blast search on amino acid sequences revealed that the protein showed similarity only with lectins from the marine sponge Spheciospongia vesparia . SFL caused agglutination of Vibrio alginolyticus and V. parahaemolyticus in a dose dependent manner and inhibited the growth rates of the virulent bacterial strains. Growth inhibition of V. alginolyticus and V. parahaemolyticus with SFL was not observed in the presence of d -galactose or asialo-porcine stomach mucin, suggesting that the lectin caused the agglutination through binding to the target receptor(s) on the surface of Vibrios. Thus, the marine sponge S. flexibilis could promise to be a good source of a lectin(s) that may be useful as a carbohydrate probe and an antibacterial reagent. [ABSTRACT FROM AUTHOR]

Three isolectins from cultivated Eucheuma denticulatum were isolated. They were commonly monomeric proteins of about 28 kDa with a range of averaged molecular weights from 27,834 to 27,868 Da among the isolectins and shared almost the same 20 N-terminal amino acid sequences. Complementary DNA (cDNA) cloning based on the rapid amplification cDNA ends (RACE) methods elucidated the full-length sequence of EDA-2 which encodes 269 amino acids, including initiating methionine, with four tandemly repeated domains of about 67 amino acids. The primary structure of EDA-2 is highly similar to those of the high-mannose N-glycan specific lectins including Oscillatoria agardhii (OAA) and Burkholderia oklahomensis EO147 (BOA) from cyanobacteria, Myxococcus xanthus (MBHA) and Pseudomonas fluorescens Pf0-1 (PFL) from bacteria, and ESA-2 from a macro red alga. The hemagglutination activities were commonly inhibited by the glycoproteins bearing high-mannose N-glycans, but not by monosaccharides examined, including mannose. In a direct binding experiment with pyridylaminated oligosaccharides, an isolectin EDA-2 exclusively bound to high-mannose type N-glycans, but not to other glycans that include complex types and a core pentasaccharide of N-glycans, indicating that it recognized the branched oligomannoside moiety. Its binding activity was subtly different among the oligomannoside structures examined, showing that the lectin has preference affinity for high-mannose type N-glycans with an exposed (α1-3) mannose residue in the D2 arm. Interestingly, EDAs, the mixture of three isolectins inhibited the growth of shrimp pathogenic bacterium, Vibrio alginolyticus, although it did not affect the growth of V. parahaemolyticus and V. harveyi. Growth inhibition of V. alginolyticus with EDAs was not observed in the presence of yeast mannan bearing high-mannose N-glycans, suggesting that EDAs caused the activity through binding to the target receptor(s) on the surface of V. alginolyticus. These results indicate that cultivated carrageenophyte E. denticulatum is a good source of a lectin(s) that may be useful as a carbohydrate probe and an antibacterial reagent. [ABSTRACT FROM AUTHOR]