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Carbene Footprinting Reveals Binding Interfaces of a Multimeric Membrane‐Spanning Protein
Author(s) -
Manzi Lucio,
Barrow Andrew S.,
Hopper Jonathan T. S.,
Kaminska Renata,
Kleanthous Colin,
Robinson Carol V.,
Moses John E.,
Oldham Neil J.
Publication year - 2017
Publication title -
angewandte chemie international edition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.831
H-Index - 550
eISSN - 1521-3773
pISSN - 1433-7851
DOI - 10.1002/anie.201708254
Subject(s) - footprinting , trimer , diazirine , micelle , chemistry , membrane , biophysics , membrane protein , protein–protein interaction , biochemistry , biology , dimer , gene , transcription factor , organic chemistry , aqueous solution
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.