Glycobiology Advance Access published online on January 26, 2007
Glycobiology, doi:10.1093/glycob/cwm007
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Strict specificity for high mannose-type N-glycans and primary structure of a red alga Eucheuma serra lectin
2 Graduate School of Biosphere Science, Hiroshima University, Kagamiyama 1-4-4, Higashi-Hiroshima 739-8528, Japan
3 Marine Greens Laboratory Co., Mori, Iyo 799-3125, Japan
4 National Research Institute of Brewing, Kagamiyama 3-7-1, Higashi-Hiroshima 739-0046, Japan
5 Sumika Chemical Analysis service, Ltd., Konohanaku 3-1-1353, Osaka 554-0022, Japan
6 Yamaki Co., Kominato, Iyo 799-3194, Japan
7 Otsuka Food Co., Chiyodaku, Tokyo 101-0053, Japan
1 Author to whom correspondence should be sent: Dr. Kanji Hori: Graduate School of Biosphere Science, Hiroshima University, Kagamiyama 1-4-4, Higashi-Hiroshima 739-8528, Japan Phone: +81-82-424-7931 Fax: +81-82-424-7916 E-mail: kanhori{at}hiroshima-u.ac.jp
Received on September 12, 2006; revised on January 14, 2007; accepted on January 16, 2007
We have elucidated the carbohydrate-binding profile of a non-monosaccharide binding lectin, named ESA-2, from the red alga Eucheuma serra by means of a lectin-immobilized column and a centrifugal ultrafiltration-HPLC method with a variety of fluorescence-labeled oligosaccharides. In both methods ESA-2 exclusively bound to high-mannose type (HM) N-glycans but not to any of other N-glycans including complex type, hybrid type and core pentasaccharides, and oligosaccharides from glycolipids. These findings indicate that ESA-2 recognizes the branched oligomannosides of the N-glycans. However, ESA-2 did not bind to any free oligomannoses examined that are constituents of the branched oligomannosides implying that the portion of the core GlcNAc residue(s) of the N-glycans is also essential for binding. Thus the algal lectin was strictly specific for HM N-glycans and recognized the extended carbohydrate structure with a minimum size of the tetrasaccharide, Man(
1-3)Man(1-6)Man(ß1-4)GlcNAc(ß1-. Kinetic analysis of binding with a HM heptasaccharide (M5) showed that ESA-2 has four carbohydrate-binding sites per polypeptide with a high association constant of 1.6 x 108 M-1. Sequence analysis, by a combination of Edman degradation and mass analyses of the intact protein and of peptides produced by its enzymic digestions, showed that ESA-2 is composed of 268 amino acids (MW 27950) with four tandemly repeated domains of 67 amino acids. The number of the repeats coincided with the number of carbohydrate-binding sites in the monomeric molecule. Surprisingly, the marine algal lectin was homologous to hemagglutinin from the soil bacterium Myxococcus xanthus.
Key words: algal lectin / amino acid sequence / Eucheuma / high mannose N-glycan specificity / Myxococcus xanthus hemagglutinin