Carbene Footprinting Reveals Binding Interfaces of a Multimeric Membrane-Spanning Protein.
Manzi L., Barrow AS., Hopper JTS., Kaminska R., Kleanthous C., Robinson CV., Moses JE., Oldham NJ.
Mapping the interaction sites between membrane-spanning proteins is a key challenge in structural biology. In this study a carbene-footprinting approach was developed and applied to identify the interfacial sites of a trimeric, integral membrane protein, OmpF, solubilised in micelles. The diazirine-based footprinting probe is effectively sequestered by, and incorporated into, the micelles, thus leading to efficient labelling of the membrane-spanning regions of the protein upon irradiation at 349 nm. Areas associated with protein-protein interactions between the trimer subunits remained unlabelled, thus revealing their location.