Strict specificity for high-mannose type N-glycans and primary structure of a red alga Eucheuma serra lectin.
We have elucidated the carbohydrate-binding profile of a non-monosaccharide-binding lectin named Eucheuma serra lectin (ESA)-2 from the red alga Eucheuma serra using a lectin-immobilized column and a centrifugal ultrafiltration-high performance liquid chromatography method with a variety of fluorescence-labeled oligosaccharides. In both methods, ESA-2 exclusively bound with high-mannose type (HM) N-glycans, but not with 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 with any of the free oligomannoses examined that are constituents of the branched oligomannosides implying that the portion of the core N-acetyl-D-glucosamine (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 pentasaccharide, Man(alpha1-3)Man(alpha1-6)Man(beta1-4)GlcNAc(beta1-4) GlcNAc. 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 1.6x10(8) 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 (molecular weight 27950) with four tandemly repeated domains of 67 amino acids. The number of 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.
Hori
Kanji
K
Graduate School of Biosphere Science, Hiroshima University, Kagamiyama 1-4-4, Higashi-Hiroshima 739-8528, Japan. kanhori@hiroshima-u.ac.jp
Sato
Yuichiro
Y
Ito
Kaori
K
Fujiwara
Yoshifumi
Y
Iwamoto
Yasumasa
Y
Makino
Hiroyuki
H
Kawakubo
Akihiro
A
D020418:Algal Proteins / Q000737:chemistry* / Q000235:genetics; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics; D002240:Carbohydrate Sequence; D008358:Mannose / Q000737:chemistry*; D037241:Mannose-Binding Lectins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D016941:Myxococcus xanthus / Q000737:chemistry / Q000235:genetics; D011134:Polysaccharides / Q000737:chemistry*; D011485:Protein Binding; D000461:Rhodophyta / Q000737:chemistry* / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D020418:Algal Proteins; D001426:Bacterial Proteins; D037241:Mannose-Binding Lectins; D011134:Polysaccharides; D008358:Mannose