ICOA UMR CRNS 7311, Universite d'Orléans, Rue de Chartres, BP 6759, 45067 Orléans Cedex 2, France.
GLYcoDiag, 2 Rue du Cristal, 45100 Orléans, France.
Int J Mol Sci. 2024 Apr 28;25(9):4826. doi: 10.3390/ijms25094826.
In the absence of naturally available galactofuranose-specific lectin, we report herein the bioengineering of GalNeoLect, from the first cloned wild-type galactofuranosidase ( sp. strain JHA19), which recognises and binds a single monosaccharide that is only related to nonmammalian species, usually pathogenic microorganisms. We kinetically characterised the GalNeoLect to confirm attenuation of hydrolytic activity and used competitive inhibition assay, with close structural analogues of Gal, to show that it conserved interaction with its original substrate. We synthetised the bovine serum albumin-based neoglycoprotein (GalNGP), carrying the multivalent Gal units, as a suitable ligand and high-avidity system for the recognition of GalNeoLect which we successfully tested directly with the galactomannan spores of (ATCC 16404). Altogether, our results indicate that GalNeoLect has the necessary versatility and plasticity to be used in both research and diagnostic lectin-based applications.
在缺乏天然存在的半乳糖呋喃糖特异性凝集素的情况下，我们在此报告了 GalNeoLect 的生物工程，它来自第一个克隆的野生型半乳糖呋喃糖苷酶（sp. strain JHA19），能够识别和结合仅与非哺乳动物物种相关的单糖，通常是致病微生物。我们对 GalNeoLect 的动力学特性进行了表征，以确认水解活性的减弱，并使用与 Gal 具有密切结构类似物的竞争性抑制测定法，表明它与原始底物保持相互作用。我们合成了基于牛血清白蛋白的糖基化蛋白（GalNGP），携带多价 Gal 单元，作为 GalNeoLect 的合适配体和高亲和力系统，我们成功地直接用（ATCC 16404）的半乳甘露聚糖孢子进行了测试。总的来说，我们的结果表明 GalNeoLect 具有必要的多功能性和可塑性，可用于研究和基于诊断凝集素的应用。
