Ogata Makoto, Anazawa Remi, Connolly Lisa, Ogata Tomomi, Wada Yuto, Tanaka Yuki, Koizumi Ami, Asano Mitsuki, Kono Hiroyuki
Faculty of Food and Agricultural Sciences, Fukushima University, 1 Kanayagawa, Fukushima, Fukushima 960-1296, Japan.
Institute of Fermentation Sciences, Fukushima University, 1 Kanayagawa, Fukushima, Fukushima 960-1296, Japan.
ACS Appl Mater Interfaces. 2024 Oct 7;16(41):56031-40. doi: 10.1021/acsami.4c08821.
Carboxymethyl cellulose (CMC) is a water-soluble cellulose derivative that is nontoxic, biocompatible, biodegradable, and nonallergenic. As developing an adsorbent material for carbohydrate-binding proteins is challenging, we aimed to synthesize CMC nanohydrogel particles (CMCGPs) with an extremely high lectin adsorption tendency in this study. CMCGPs were used as the backbone of an adsorption carrier that was synthesized by cross-linking CMC with ethylene glycol diglycidyl ether. A series of glycoside-immobilized CMCGPs were synthesized by binding two types of glycans (LacNAc and lactose) to the polyvalent carboxymethyl groups that are present on the CMCGP surface and act as reaction sites. These immobilized glycosides function as molecular recognition sites. Glycan moieties were incorporated into the CMCGP backbone at degrees of immobilization (DI) ranging from 8.7 to 21.0% by altering the reaction composition. LacNAc-CMCGP () showed a 19.9% DI of LacNAc glycoside to the CMCGP carboxymethyl group; on average, its particle size swelled to 418 nm in phosphate-buffered saline, which is approximately 1.4 times its dry-state size. Analyzing the adsorbent properties of glyco-CMCGPs using a lectin-binding assay showed the high structural specificity of glyco-CMCGPs to lectins. The equilibrium isotherm data was explained by the Langmuir adsorption model. Notably, compound adsorbed 1.95 ± 0.05 μg of wheat germ agglutinin (WGA) lectin per 1.0 μg-dry of particles at an adsorption equilibrium time of a few minutes. Furthermore, solid-state C nuclear magnetic resonance analysis showed that WGA lectin retained its natural structure without denaturation after binding to LacNAc-CMCGP. These results were also supported by affinity purification experiments of WGA from raw wheat germ extract using LacNAc-CMCGP, demonstrating that glyco-CMCGP is capable of adsorbing and desorbing lectin while maintaining its biological activity. Thus, multivalent glycoside-immobilized CMCGPs that use woody biomass derivatives as the backbone are expected to be applied as biorefinery materials, which specifically and abundantly adsorb not just plant lectins but also pathogenic viruses and toxin proteins.
羧甲基纤维素（CMC）是一种水溶性纤维素衍生物，无毒、具有生物相容性、可生物降解且无致敏性。由于开发用于碳水化合物结合蛋白的吸附材料具有挑战性，因此在本研究中，我们旨在合成具有极高凝集素吸附倾向的CMC纳米水凝胶颗粒（CMCGPs）。CMCGPs用作吸附载体的骨架，该吸附载体是通过将CMC与乙二醇二缩水甘油醚交联合成的。通过将两种类型的聚糖（N-乙酰乳糖胺和乳糖）结合到CMCGP表面存在的多价羧甲基基团上并将其作为反应位点，合成了一系列固定化糖苷的CMCGPs。这些固定化糖苷用作分子识别位点。通过改变反应组成，将聚糖部分以8.7%至21.0%的固定化度（DI）掺入CMCGP骨架中。N-乙酰乳糖胺-CMCGP（）显示N-乙酰乳糖胺糖苷与CMCGP羧甲基基团的DI为19.9%；在磷酸盐缓冲盐水中，其平均粒径膨胀至418 nm，约为其干态尺寸的1.4倍。使用凝集素结合试验分析糖基化CMCGPs的吸附特性，结果表明糖基化CMCGPs对凝集素具有高度的结构特异性。平衡等温线数据可用朗缪尔吸附模型解释。值得注意的是，在几分钟的吸附平衡时间下，化合物每1.0 μg干颗粒吸附1.95±0.05 μg的麦胚凝集素（WGA）凝集素。此外，固态碳核磁共振分析表明，WGA凝集素与N-乙酰乳糖胺-CMCGP结合后保持其天然结构而未变性。使用N-乙酰乳糖胺-CMCGP从生麦胚提取物中对WGA进行亲和纯化实验也支持了这些结果，表明糖基化CMCGP能够吸附和解吸凝集素，同时保持其生物活性。因此，以木质生物质衍生物为骨架的多价固定化糖苷CMCGPs有望用作生物精炼材料，其不仅能特异性且大量地吸附植物凝集素，还能吸附致病病毒和毒素蛋白。
