Mathias Winterhalter, Wanling Song, Hualiang Jiang, Yechun Xu, Harsha Bajaj, Chady Nasrallah, Jacques-Philippe Colletier, Laboratory of Receptor Research, Shanghai Institute of Materia Medica, School of Engineering and Science [Bremen] (JU-SES), Jacobs University [Bremen], Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), and Thomas, Frank more...
Bacterial porins are water-filled β-barrel channels that allow translocation of solutes across the outer membrane. They feature a constriction zone, contributed by the plunging of extracellular loop 3 (L3) into the channel lumen. Porins are generally in the open state, but undergo gating in response to external voltages. To date the underlying mechanism is unclear. Here we report results from molecular dynamics simulations on the two porins of Providenica stuartii, Omp-Pst1 and Omp-Pst2, which display distinct voltage sensitivities. Voltage gating was observed in Omp-Pst2, where the binding of cations in-between L3 and the barrel wall results in exposing a conserved aromatic residue in the channel lumen, thereby halting ion permeation. Comparison of Omp-Pst1 and Omp-Pst2 structures and trajectories suggests that their sensitivity to voltage is encoded in the hydrogen-bonding network anchoring L3 onto the barrel wall, as we observed that it is the strength of this network that governs the probability of cations binding behind L3. That Omp-Pst2 gating is observed only when ions flow against the electrostatic potential gradient of the channel furthermore suggests a possible role for this porin in the regulation of charge distribution across the outer membrane and bacterial homeostasis., Author Summary Porins are the main conduits for hydrophilic nutrients and ions uptake into the periplasm of Gram-negative bacteria. Their translocation permeability is determined by the amino-acid distribution on their extracellular loop L3. Bacterial porin channels have long been known to undergo step-wise gating, under the application of a transmembrane potential. Yet the exact molecular mechanism by which gating is achieved and the exact relevance of this evolved characteristic remain elusive. In the present study, we report on electrophysiology experiments and molecular dynamics simulations on the two general-diffusion porins of Providencia stuartii, Omp-Pst1 and Omp-Pst2. Our results show that gating in Omp-Pst2 occurs as the result of L3 displacement, which follows from the binding of cations in acidic niches between L3 and the barrel wall and effects in exposing the side chain of a highly conserved aromatic residue at the tip of L3 in the channel lumen. That Omp-Pst2 displays asymmetric voltage sensitivity and that the likelihood of gating is increased when cations transit from the extracellular to the intracellular side suggests voltage-gating underlies a regulatory role in bacterial homeostasis. Rational antibiotic-design strategies based on the maximization of antibiotic penetration and accumulation at their target sites, should take this role into account. more...