Faculty of Food and Agricultural Sciences, Fukushima University, 1 Kanayagawa, Fukushima, Fukushima, 960-1296, Japan; Department of Applied Chemistry and Biochemistry, National Institute of Technology, Fukushima College, 30 Nagao, Iwaki, Fukushima, 970-8034, Japan.
Department of Applied Chemistry and Biochemistry, National Institute of Technology, Fukushima College, 30 Nagao, Iwaki, Fukushima, 970-8034, Japan.
Carbohydr Res. 2022 Sep;519:108624. doi: 10.1016/j.carres.2022.108624. Epub 2022 Jun 16.
Glycopolypeptide-immobilized particulates exhibit high binding selectivities and affinities for several analytes. However, to date, the conditions for the synthesis of glycopolypeptide-immobilized particulates have not been optimized and the application of these particulates as carriers for affinity chromatography has not been reported. Accordingly, herein, as a model compound for determining the optimal conditions for the immobilization of an artificial glycopolymer on hexyl-containing hybrid silica particulates (HSPs), the glycopolypeptide poly [GlcNAcβ1,4GlcNAc-β-NHCO-(CH)NH-/CH(CH)NH-/γ-PGA] (3) containing multivalent chitobiose moieties and multivalent decyl groups with a γ-polyglutamic acid backbone was synthesized. Immobilization of 3 on HSPs under each condition was evaluated by a lectin-binding assay using wheat germ (Triticum vulgaris) agglutinin (WGA), which is an N-acetylglucosamine-binding lectin. As a result, the optimal immobilization conditions for HSPs at 25 mg/mL were obtained at dimethyl sulfoxide (DMSO) concentration of reaction solvent in the range of 1:9 to 4:6 and a compound 3 concentration in the range of 125 nM-1250 nM. Furthermore, the influence of the alkyl group structure introduced into glycopolypeptide for imparting hydrophobicity to it on the immobilization of glycopolypeptide on HSPs was investigated. As a result of comparing three types, poly [GlcNAcβ1,4GlcNAc-β-NHCO-(CH)NH-/γ-PGA] (1) with no alkyl group, poly [GlcNAcβ1,4GlcNAc-β-NHCO-(CH)NH-/CH(CH)NH-/γ-PGA] (2) with a pentyl group, and 3 with a decyl group, 3 showed the best immobilization efficiency on HSPs. Finally, 1 mg 3-immobilized HSPs prepared under the optimum conditions adsorbed approximately 7.5 μg WGA in a structure-specific manner. We also achieved a simple WGA purification from raw wheat germ extract as a practical example using 3-immobilized HSPs. We believe that in the future, these glycopolypeptide-immobilized particulates will be used not only for the purification of plant lectins, but also as specific adsorbents for various lectins-like substances such as in vivo lectins, pathogenic viruses, and toxin proteins.
糖肽固定颗粒对多种分析物表现出高的结合选择性和亲和力。然而，迄今为止，糖肽固定颗粒的合成条件尚未优化，并且这些颗粒作为亲和层析载体的应用尚未报道。因此，本文以含有多价壳二糖结构单元和多价癸基的糖肽聚[GlcNAcβ1,4GlcNAc-β-NHCO-(CH)NH-/CH(CH)NH-/γ-PGA]（3）为模型化合物，用于确定在含己基的杂化硅胶颗粒（HSPs）上固定人工糖聚合物的最佳条件，该糖肽含有多价壳二糖结构单元和多价癸基，具有γ-聚谷氨酸主链。通过使用麦胚凝集素（WGA）进行凝集素结合试验评估 3 在 HSPs 上的固定情况，WGA 是一种 N-乙酰葡萄糖胺结合凝集素。结果表明，当反应溶剂中二甲基亚砜（DMSO）浓度在 1:9 至 4:6 范围内，化合物 3 浓度在 125 nM-1250 nM 范围内时，HSPs 的最佳固定条件为 25 mg/mL。此外，还研究了引入糖肽中的烷基结构对赋予其疏水性的影响，以研究其对糖肽在 HSPs 上固定的影响。通过比较三种类型，即无烷基的聚[GlcNAcβ1,4GlcNAc-β-NHCO-(CH)NH-/γ-PGA]（1）、具有戊基的聚[GlcNAcβ1,4GlcNAc-β-NHCO-(CH)NH-/CH(CH)NH-/γ-PGA]（2）和具有癸基的 3，发现 3 对 HSPs 的固定效率最好。最后，在最佳条件下制备的 1mg 3-固定 HSPs 以结构特异性的方式吸附了约 7.5μg 的 WGA。我们还使用 3-固定的 HSPs 从原始小麦胚提取物中成功地实现了 WGA 的简单纯化，作为一个实际的例子。我们相信，在未来，这些糖肽固定颗粒不仅将用于植物凝集素的纯化，还将用于各种类似凝集素的物质，如体内凝集素、致病病毒和毒素蛋白的特异性吸附剂。
