Strict specificity for high-mannose type N-glycans and primary structure of a red alga Eucheuma serra lectin
Strict specificity for high-mannose type N-glycans and primary structure of a red alga Eucheuma serra lectin
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DOI：
10.1093/glycob/cwm007
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发表时间：
2007-05-01
期刊：
影响因子：
4.3
通讯作者：
Kawakubo, Akihiro
中科院分区：
文献类型：
--
作者：
Hori, Kanji;Sato, Yuichiro;Kawakubo, Akihiro
采用凝集素固定化柱和离心超滤-高效液相色谱法，对红藻真毛凝集素（Eucheuma serra lectin， ESA）-2的碳水化合物结合谱进行了分析。在这两种方法中，ESA-2只与高甘露糖型（HM） n -聚糖结合，而不与任何其他n -聚糖结合，包括复合型、杂化型和核心五糖，以及来自糖脂的低聚糖。这些发现表明ESA-2能够识别n -聚糖的支链寡聚糖。然而，ESA-2没有与任何被检测的游离寡糖结合，这些寡糖是支链寡糖苷的组成部分，这意味着n -聚糖的核心n -乙酰-d-氨基葡萄糖（GlcNAc）残基部分也是结合所必需的。因此，海藻凝集素对HM n -聚糖具有严格的特异性，并识别出最小尺寸的扩展碳水化合物结构Man(α 1-3)Man(α 1-6)Man(β 1-4)GlcNAc(β 1-4)GlcNAc。与HM七糖（M5）结合的动力学分析表明，ESA-2每个多肽具有4个碳水化合物结合位点，其高结合常数为1.6 x 10(8) M-1。通过对完整蛋白和酶解产生的肽的Edman降解和质量分析，序列分析表明ESA-2由268个氨基酸（分子量27950）组成，其中有67个氨基酸的4个连续重复结构域。重复次数与单分子中碳水化合物结合位点的数量一致。令人惊讶的是，这种海藻凝集素与土壤细菌黄粘球菌的血凝素是同源的。
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(alpha 1-3)Man(alpha 1-6)Man(beta 1-4)GlcNAc(beta 1-4) 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 1.6 x 10(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.