Stourman Nina V
Department of Chemical and Biological Sciences, Youngstown State University, Youngstown, OH, USA.
Methods Mol Biol. 2025;2917:133-141. doi: 10.1007/978-1-0716-4478-2_12.
Fluorogenic sugar derivatives are useful substrates for the detection of glycosidase activity within a native polyacrylamide gel. When an enzyme cleaves the glycosidic bond, a fluorescent compound is released at the protein's location, forming a band visible under UV light. This method can be used to identify proteins with glycosidase activity within a mixture, evaluate the effect of mutations on enzyme activity, and even provide information about the quaternary structure of a glycosidase. Here, we present a protocol for glycosidase zymography and demonstrate its utility as a tool to probe the active oligomeric form of a novel bacterial β-galactosidase.
荧光糖衍生物是用于检测天然聚丙烯酰胺凝胶内糖苷酶活性的有用底物。当酶切割糖苷键时，荧光化合物在蛋白质所在位置释放，形成在紫外光下可见的条带。该方法可用于鉴定混合物中具有糖苷酶活性的蛋白质，评估突变对酶活性的影响，甚至提供有关糖苷酶四级结构的信息。在这里，我们展示了一种糖苷酶酶谱分析方法，并证明了其作为探测新型细菌β-半乳糖苷酶活性寡聚体形式工具的实用性。
