Department of Chemistry, Brown University, Providence, Rhode Island 02912, United States.
Division of Plant Science and Technology, Interdisciplinary Plant Group, The Missouri Maize Center, University of Missouri, Columbia, Missouri 65211, United States.
Bioconjug Chem. 2023 Aug 16;34(8):1398-1406. doi: 10.1021/acs.bioconjchem.3c00184. Epub 2023 Aug 3.
Small molecule fluorescent probes that bind selectively to plant cell wall polysaccharides have been instrumental in elucidating the localization and function of these glycans. Arabinogalactan proteins (AGPs) are cell wall proteoglycans implicated in essential functions such as cell signaling, plant growth, and programmed cell death. There is currently no small molecule probe capable of fluorescently labeling AGPs. The Yariv reagents are the only small molecules that bind AGPs, and have been used to study AGP function and isolate AGPs via precipitation of an AGP-Yariv complex. However, the Yariv reagents are not fluorescent, rendering them ineffective for localization studies using fluorescence microscopy. A fluorescent version of a Yariv reagent that is capable of both binding as well as imaging AGPs would provide a powerful tool for studying AGPs in planta. Herein, we describe the synthesis of an azido analog of the Yariv reagent that can be further functionalized with a fluorophore to provide a glycoconjugate that binds AGPs and is fluorescent. We show that the modified reagent binds gum arabic in in vitro binding assays when used in conjunction with the βGlcYariv reagent. Fluorescent imaging of AGPs in fixed maize leaf tissue enables localization of AGPs to cell walls in the leaf. Significantly, imaging can also be carried out using fresh tissue. This represents the first small molecule probe that can be used to visualize AGPs using fluorescence microscopy.
能够选择性结合植物细胞壁多糖的小分子荧光探针对于阐明这些糖的定位和功能至关重要。阿拉伯半乳聚糖蛋白 (AGP) 是一种细胞壁糖蛋白，参与细胞信号转导、植物生长和程序性细胞死亡等基本功能。目前还没有能够荧光标记 AGP 的小分子探针。Yariv 试剂是唯一能够结合 AGP 的小分子，已被用于研究 AGP 功能，并通过沉淀 AGP-Yariv 复合物来分离 AGP。然而，Yariv 试剂本身不具有荧光性，因此无法用于荧光显微镜下的定位研究。一种能够结合和成像 AGP 的 Yariv 试剂的荧光版本将为在体内研究 AGP 提供一种强大的工具。在此，我们描述了 Yariv 试剂的叠氮类似物的合成，该类似物可以进一步与荧光团官能化，提供一种结合 AGP 且具有荧光性的糖缀合物。我们表明，当与βGlcYariv 试剂一起使用时，该修饰的试剂在体外结合测定中结合阿拉伯胶。固定的玉米叶片组织中 AGP 的荧光成像使 AGP 能够定位于叶片细胞壁。重要的是，也可以使用新鲜组织进行成像。这代表了第一个可以使用荧光显微镜可视化 AGP 的小分子探针。
