Abstract:
Peripheral membrane proteins participate in numerous biological pathways. Thus, methods to analyze their membrane‐binding characteristics have become important. In this report, we detail protocols for the synthesis and utilization of a photoactivable fluorescent lipid as a reporter to monitor membrane binding of proteins. The assay, referred to as proximity‐based labeling of membrane‐associated proteins (PLiMAP), is based on UV activation of a fluorescent lipid reporter, which in turn crosslinks with proteins bound to membranes and renders them fluorescent. © 2020 Wiley Periodicals LLC.Basic Protocol 1: Synthesis of BODIPY‐diazirine phosphatidylethanolamine (BDPE) Basic Protocol 2: Preparation of BDPE‐containing liposomes Basic Protocol 3: Performing PLiMAP with a candidate protein Basic Protocol 4: Quantitation of liposome‐binding properties of the candidate protein from analyzing in‐gel fluorescence Support Protocol: Purification of GST‐2×P4M domain of SidM protein