Department of Chemistry, University of Connecticut, 55 N. Eagleville Road U3060, Storrs, CT 06269, USA.
Laboratory of Biochemistry, Institute Químic de Sarrià, University Ramon Llull, 08017 Barcelona, Spain.
Int J Mol Sci. 2021 Apr 26;22(9):4497. doi: 10.3390/ijms22094497.
Building-up and breaking-down of carbohydrates are processes common to all forms of life. Glycoside hydrolases are a broad class of enzymes that play a central role in the cleavage of glycosidic bonds, which is fundamental to carbohydrate degradation. The large majority of substrates are five- and six-membered ring glycosides. Our interest in seven-membered ring septanose sugars has inspired the development of a way to search for septanoside hydrolase activity. Described here is a strategy for the discovery of septanoside hydrolases that uses synthetic indolyl septanosides as chromogenic substrates. Access to these tool compounds was enabled by a route where septanosyl halides act as glycosyl donors for the synthesis of the indolyl septanosides. The screening strategy leverages the known dimerization of 3-hydroxy-indoles to make colored dyes, as occurs when the β-galactosidase substrate X-Gal is hydrolyzed. Because screens in bacterial cells would enable searches in organisms that utilize heptoses or from metagenomics libraries, we also demonstrate that septanosides are capable of entering cells through the use of a BODIPY-labeled septanoside. The modularity of the indolyl septanoside synthesis should allow the screening of a variety of substrates that mimic natural structures via this general approach.
碳水化合物的合成与分解是所有生命形式共有的过程。糖苷水解酶是一大类酶，在糖苷键的断裂中发挥着核心作用，而糖苷键的断裂是碳水化合物降解的基础。绝大多数的底物是五元和六元环糖苷。我们对七元环庚酮糖的兴趣激发了寻找庚酮糖苷水解酶活性的方法。本文描述了一种使用合成吲哚基庚酮糖苷作为显色底物来发现庚酮糖苷水解酶的策略。这些工具化合物的获得得益于一种方法，其中庚糖卤化物作为吲哚基庚酮糖苷合成的糖基供体。该筛选策略利用了 3-羟基吲哚的已知二聚化作用来产生有色染料，就像β-半乳糖苷酶底物 X-Gal 被水解时发生的那样。因为在细菌细胞中的筛选可以在利用庚糖或来自宏基因组文库的生物体中进行搜索，所以我们还证明了通过使用 BODIPY 标记的庚酮糖苷，庚酮糖苷能够进入细胞。吲哚基庚酮糖苷的合成的模块化应该允许通过这种通用方法筛选各种模拟天然结构的底物。
