基于响应型蛋白标签配体的FRET生物传感器用于碱性磷酸酶活性检测

Fluorescent protein-based FRET biosensors can monitor the spatiotemporal changes of molecules in real time and are important tools for elucidating signal transduction in living cells. However, most of these sensors rely on the change of the distance between the donor and the acceptor caused by the domain, so they need to be reconstructed by protein engineering technology when detecting different targets. In order to simplify the sensor design steps, Fluorescence-Activating and absorption-Shifted Tag (FAST) were introduced into the green fluorescent protein (GFP)-based FRET sensor, and the activity detection of alkaline phosphatase (ALP) was realized by modifying the phosphate group on the ligand molecule. The sensor has a specific and sensitive response to ALP, and successfully detected ALP in cell lysates. The successful construction of such sensors expands the design ideas of protein-based FRET sensors and lays the foundation for the specific detection of various enzymes in living cells.