Riera Roger, Hogervorst Tim P, Doelman Ward, Ni Yan, Pujals Silvia, Bolli Evangelia, Codée Jeroen D C, van Kasteren Sander I, Albertazzi Lorenzo
Department of Biomedical Engineering, Institute for Complex Molecular Systems, Eindhoven University of Technology, Eindhoven, the Netherlands.
Department of Bio-Organic Synthesis, Leiden Institute of Chemistry, Leiden University, Leiden, the Netherlands.
Nat Chem Biol. 2021 Dec;17(12):1281-1288. doi: 10.1038/s41589-021-00896-2. Epub 2021 Nov 11.
Most lectins bind carbohydrate ligands with relatively low affinity, making the identification of optimal ligands challenging. Here we introduce a point accumulation in nanoscale topography (PAINT) super-resolution microscopy method to capture weak glycan-lectin interactions at the single-molecule level in living cells (Glyco-PAINT). Glyco-PAINT exploits weak and reversible sugar binding to directly achieve single-molecule detection and quantification in cells and is used to establish the relative k and k rates of a synthesized library of carbohydrate-based probes, as well as the diffusion coefficient of the receptor-sugar complex. Uptake of ligands correlates with their binding affinity and residence time to establish structure-function relations for various synthetic glycans. We reveal how sugar multivalency and presentation geometry can be optimized for binding and internalization. Overall, Glyco-PAINT represents a powerful approach to study weak glycan-lectin interactions on the surface of living cells, one that can be potentially extended to a variety of lectin-sugar interactions.
大多数凝集素与碳水化合物配体的结合亲和力相对较低，这使得鉴定最佳配体具有挑战性。在此，我们引入一种纳米级形貌点积累（PAINT）超分辨率显微镜方法，以在活细胞的单分子水平上捕获弱聚糖-凝集素相互作用（糖-PAINT）。糖-PAINT利用弱且可逆的糖结合直接在细胞中实现单分子检测和定量，并用于确定合成的基于碳水化合物的探针文库的相对k和k速率，以及受体-糖复合物的扩散系数。配体的摄取与其结合亲和力和停留时间相关，从而为各种合成聚糖建立结构-功能关系。我们揭示了如何优化糖的多价性和呈现几何结构以实现结合和内化。总体而言，糖-PAINT是研究活细胞表面弱聚糖-凝集素相互作用的有力方法，并且有可能扩展到各种凝集素-糖相互作用。
