Live Cell Imaging and Profiling of Cysteine Cathepsin Activity Using a Quenched Activity-Based Probe
Live Cell Imaging and Profiling of Cysteine Cathepsin Activity Using a Quenched Activity-Based Probe
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DOI：
10.1007/978-1-4939-6439-0_11
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发表时间：
2017-01-01
期刊：
影响因子：
--
通讯作者：
Verdoes, Martijn
中科院分区：
文献类型：
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作者：
Edgington-Mitchell, Laura E.;Bogyo, Matthew;Verdoes, Martijn
由于蛋白酶活性受到结构和环境影响的高度调节，蛋白酶的丰度通常与其活性不直接相关。因为在大多数情况下，是蛋白酶的活性引起了它的生物学相关性，因此报告这种活性的工具对研究界具有很大的价值。基于活性的探针（ABPs）是一种小分子工具，可用于监测和分析复杂生物系统中的蛋白酶活性。荧光猝灭ABPs （qABPs）本质上是“暗”的，只在与目标蛋白酶反应后才发出荧光，非常适合小动物无创荧光成像和活细胞荧光显微镜等成像技术。qABPs的另一个强大特性是它们对标记蛋白酶的共价和不可逆修饰，从而能够深入表征靶标。本文描述了一种泛半胱氨酸组织蛋白酶qABP BMV109的合成，并将该探针应用于活细胞荧光成像和荧光SDS-PAGE半胱氨酸组织蛋白酶活性分析。
Since protease activity is highly regulated by structural and environmental influences, the abundance of a protease often does not directly correlate with its activity. Because in most of the cases it is the activity of a protease that gives rise to its biological relevance, tools to report on this activity are of great value to the research community. Activity-based probes (ABPs) are small molecule tools that allow for the monitoring and profiling of protease activities in complex biological systems. The class of fluorescent quenched ABPs (qABPs), being intrinsically "dark" and only emitting fluorescence after reaction with the target protease, are ideally suited for imaging techniques such as small animal noninvasive fluorescence imaging and live cell fluorescence microscopy. An additional powerful characteristic of qABPs is their covalent and irreversible modification of the labeled protease, enabling in-depth target characterization. Here we describe the synthesis of a pan-cysteine cathepsin qABP BMV109 and the application of this probe to live cell fluorescence imaging and fluorescent SDS-PAGE cysteine cathepsin activity profiling.