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Small‐Molecule Permeation across Membrane Channels: Chemical Modification to Quantify Transport across OmpF
Author(s) -
Wang Jiajun,
Bafna Jayesh Arun,
Bhamidimarri Satya Prathyusha,
Winterhalter Mathias
Publication year - 2019
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.201814489
Subject(s) - permeation , chemistry , membrane , cysteine , biophysics , molecule , covalent bond , small molecule , membrane transport , combinatorial chemistry , biochemistry , organic chemistry , biology , enzyme
Biological channels facilitate the exchange of molecules across membranes, but general tools to quantify transport are missing. Electrophysiology is the method of choice to study the functional properties of channels. However, analyzing the current fluctuation of channels typically does not identify successful transport, that is, distinguishing translocation from binding. To distinguish both processes, we added an additional barrier at the channel exit acting as a molecular counter. To identify permeation, we compare the molecule residence time in the native channel with one that is chemically modified at the exit. We use the well‐studied outer membrane channel from E. coli, OmpF. Position 181, which is below the constriction region, was subsequently mutated into cysteine (E181C) in an otherwise cysteine‐free system, then functionalized by covalent binding with one of the two blockers MTSES or GLT. We measured the passage of model peptides, mono‐, tri‐, hepta‐arginine and of norfloxacin, as an example for antibiotic permeation.