Rationalizing the permeation of polar antibiotics into Gram-negative bacteria

Abstract

The increasing level of antibiotic resistance in Gram-negative bacteria, together with the lack of new potential drug scaffolds in the pipeline, make the problem of infectious diseases a global challenge for modern medicine. The main reason that Gram-negative bacteria are particularly challenging is the presence of an outer cell-protecting membrane, which is not present in Gram-positive species. Such an asymmetric bilayer is a highly effective barrier for polar molecules. Several protein systems are expressed in the outer membrane to control the internal concentration of both nutrients and noxious species, in particular: (i) water-filled channels that modulate the permeation of polar molecules and ions according to concentration gradients, and (ii) efflux pumps to actively expel toxic compounds. Thus, besides expressing specific enzymes for drugs degradation, Gram-negative bacteria can also resist by modulating the influx and efflux of antibiotics, keeping the internal concentration low. However, there are no direct and robust experimental methods capable of measuring the permeability of small molecules, thus severely limiting our knowledge of the molecular mechanisms that ultimately control the permeation of antibiotics through the outer membrane. This is the innovation gap to be filled for Gram-negative bacteria. This review is focused on the permeation of small molecules through porins, considered the main path for the entry of polar antibiotics into Gram-negative bacteria. A fundamental understanding of how these proteins are able to filter small molecules is a prerequisite to design/optimize antibacterials with improved permeation. The level of sophistication of modern molecular modeling algorithms and the advances in new computer hardware has made the simulation of such complex processes possible at the molecular level. In this work we aim to share our experience and perspectives in the context of a multidisciplinary extended collaboration within the IMI-Translocation consortium. The synergistic combination of structural data, in vitro assays and computer simulations has proven to give new insights towards the identification and description of physico-chemical properties modulating permeation. Once similar general rules are identified, we believe that the use of virtual screening techniques will be very helpful in searching for new molecular scaffolds with enhanced permeation, and that molecular modeling will be of fundamental assistance to the optimization stage.

Getting Drugs into Gram-Negative Bacteria: Rational Rules for Permeation through General Porins

“…While previous studies have been focused in the exploration of the chemical space of molecules only, here we included the exploration of porin diversity via the structural determinants. This has allowed us to analyze experimental transport data in term of pore-molecule interaction terms and validate the underlying molecular mechanism 37 . The proposed scoring function is based on well-defined physicochemical properties coming from main porin features: an entropic barrier, related to the hourglass shape of porins, modulated by the electrostatic interactions of the molecule in the charge-segregated CR of the porin.…”

“…The scoring function for permeability: connecting structure to transport In order to quantify antibiotic permeability through porins, we need a theoretical model for permeability to understand how the molecular-scale properties of porins control the macroscopic flow of molecules. If we treat the antibiotic translocation process as a one-dimensional diffusion-drift problem 36,37 , the molecular flux through the channel follows Fick's law, J =-N a PS 0 Dc , where Δc is the difference of the antibiotic molar concentrations, S 0 is the geometrical cross section at the mouth of the channel and N a is the Avogadro constant. The permeability coefficient P, reflects the ability of a certain molecule to translocate through the channel.…”

Fosfomycin Permeation through the Outer Membrane Porin OmpF

“…In general, the transport of antibiotic molecules, especially across the OM of Gram-negative bacteria, has been identified as one of the key problems in antibiotics research (14)(15)(16). Recently, considerable effort-both experimentally as well as theoretically-has been put into this subject (see, for example, (17)(18)(19)(20)(21)(22)(23)(24)(25)), which is quite complex, partially because of the large diversity of channels and the so far often-incomplete knowledge of their structure and functional properties.…”

Exploring bacterial outer membrane barrier to combat bad bugs

“… 7 , 15 , 22 The method includes reconstitution of a single or multiple Omps into an artificial planar lipid bilayer and further uses transmembrane potential-driven ion current across the channel as a detection probe. 7 , 67 Using ion current as a probe specifically demonstrates very well-characterized electrophysiological properties of the Omps, 15 , 34 , 45 , 65 , 66 , 84 , 106 , 119 – 121 including size, 122 , 123 single-channel conductance, channel ion selectivity, 58 , 75 , 76 , 90 , 99 – 101 channel gating dynamics, and more. 47 , 95 , 109 Likewise, the size of Omps is a key factor defining transport through the channel.…”

“…In the current scenario, MS is well suited to obtain a particular information at an atomic scale. 121 Thus far, knowledge of the antibiotic translocation problem has pointed essentially toward three mechanisms including diffusion with molecule binding, a mechanism based on pore dehydration induced by the permeating molecule, and slow diffusion with molecule binding. 50 , 61 , 62 , 70 , 71 , 97 , 99 , 121 Further, to discriminate among these mechanisms, and to attain a better description of the Omps behavior and their role in substrate transport, understanding the communication between pore and substrate is essential.…”