PLiMAP: Proximity‐Based Labeling of Membrane‐Associated Proteins

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