Efficient Synthesis of an Alkyne Fluorophosphonate Activity‐Based Probe and Applications in Dual Colour Serine Hydrolase Labelling

Abstract

AbstractTagged fluorophosphonates (FP‐probes) were the first activity‐based probes to be reported in 1999. More than 20 years later, FP‐probes are widely used tools for the profiling of serine hydrolase‐activity. However, their synthesis tends to be cumbersome and low‐yielding. In this paper, we report a short and high‐yielding route towards an FP‐alkyne derivative. This probe is suitable for bioorthogonal 2‐step detection of serine hydrolases. Alternatively, it can be converted to an array of fluorescent probes via click chemistry to azide‐functionalized fluorophores. Here, we constructed red‐ and green‐fluorescent FP‐probes and applied these in dual colour labelling of whole proteomes. Specifically, we illustrate application in inhibitor assessment in tissue lysates and in a ‘pulse‐chase ABPP’ experiment in whole cells. Overall, we think that these probes, due to their high yield and different colours, are attractive for novel applications within ABPP.