The Optimal Permeation of Cyclic Boronates to Cross the Outer Membrane via the Porin Pathway

We investigated the diffusion of three cyclic boronates formulated as beta-lactamase inhibitors through the porin OmpF to evaluate their potential to cross OM via the porin pathway. The three nonbeta-lactam molecules diffuse through the porin eyelet region with the same mechanism observed for beta-lactam molecules and diazobicyclooctan derivatives, with the electric dipole moment aligned with the transversal electric field. In particular, the BOH group can interact with both the basic ladder and the acidic loop L3, which is characteristic of the size-constricted region of this class of porins. On one hand, we confirm that the transport of small molecules through enterobacter porins has a common general mechanism; on the other, the class of cyclic boronate molecules does not seem to have particular difficulties in diffusing through enterobacter porins, thus representing a good scaffold for new anti-infectives targeting Gram-negative bacteria research

Promising Perspectives on the Use of Fullerenes as Efficient Containers for Beryllium Atoms

The possibility of using fullerenes as containers for toxic beryllium atoms is studied by a multi-scale approach in which first-principles and classical molecular dynamics simulations are combined. By studying the energetics, electronic and spectroscopic properties of Be-fullerene systems and by simulating their interaction at finite temperature in vacuo and in representative biological environments it is concluded that: i) Be endohedral complexes can be obtained by implanting Be atoms at energies >2.3 eV that is consistent with laser implantation technologies; ii) it is in principle possible to distinguish stable endohedral complexes from metastable exohedral ones by optical absorption, suggesting that optical spectroscopy can be a valuable a non-destructive technique to assist the synthesis and the control of implanted films iii) the Be-endohedral complexes are long-lived and thermodynamically stable and can confine beryllium both in vacuo and in aqueous solution; iv) Be@C60 complexes are likely unable to penetrate the selectivity filters of a prototypical protein showing that fullerene prevents undesired interactions with biomolecules and toxicity effects of Be2+ related to replacement of the Ca2+. Overall, these results provide an assessment on the possibility to encapsulate Be atoms into fullerenes by ion implantation to synthesize inert and highly stable and safe molecular containers for toxic beryllium radionuclides. Great opportunities are expected for the realization and application of Be-C60 complexes to nanotechnology and nanomedicine with particularly appealing perspectives in the field of neutron capture therapy of cancer

Getting drugs into gram-negative bacteria: rational rules for permeation through general porins

Small, hydrophilic molecules, including most important antibiotics in clinical use, cross the Gram-negative outer membrane through the water-filled channels provided by porins. We have determined the X-ray crystal structures of the principal general porins from three species of Enterobacteriaceae, namely Enterobacter aerogenes, Enterobacter cloacae and Klebsiella pneumoniae and determined their antibiotic permeabilities as well as those of the orthologues from Escherichia coli. Starting from the structure of the porins and molecules we propose a physical mechanism underlying transport and condense it in a computationally efficient scoring function. The scoring function shows good agreement with in-vitro penetration data and will enable the screening of virtual databases to identify molecules with optimal permeability through porins and help to guide the optimization of antibiotics with poor permeation

Permeability Through Bacterial Porins Dictates Whole Cell Compound Accumulation

The lack of new drugs for Gram-negative pathogens is a global threat to modern medicine. The complexity of their cell envelope, with an additional outer membrane, hinders internal accumulation and thus, the access of molecules to targets. Our limited understanding of the molecular basis for compound influx and efflux from these pathogens is a major bottleneck for the discovery of effective antibacterial compounds. Here we analyse the correlation between the whole-cell compound accumulation of ~200 molecules and their predicted porin permeability coefficient (influx), using a recently developed scoring function. We found a strong linear relationship (75%) between the two, confirming porins key role in compound penetration. Further, the remarkable prediction ability of the scoring function demonstrates its potentiality to guide the optimization of hits to leads as well as the possibility of screening ultra-large virtual libraries. Eventually, the analysis of false positives, molecules with high-predicted influx but low accumulation, provides new hints on the molecular properties behind efflux.<br /

The Influence of Permeability through Bacterial Porins in Whole-Cell Compound Accumulation

The lack of new drugs for Gram-negative pathogens is a global threat to modern medicine. The complexity of their cell envelope, with an additional outer membrane, hinders internal accumulation and thus, the access of molecules to their targets. Our limited understanding of the molecular basis for compound influx and efflux from these pathogens is a major bottleneck for the discovery of effective antibacterial compounds. Here we analyse the correlation between the whole-cell compound accumulation of ~200 molecules and their predicted porin permeability coefficient (influx), using a recently developed scoring function. We found a strong linear relationship (74%) between the two, confirming porins key in compound uptake in Gram-negative bacteria. The analysis of this unique dataset aids to better understand the molecular descriptors behind whole-cell accumulation and molecular uptake in Gram-negative bacteria

Sensing Single Molecule Penetration into Nanopores: Pushing the Time Resolution to the Diffusion Limit

To quantify small molecule penetration into and eventually permeation through nanopores, we applied an improved excess-noise analysis of the ion current fluctuation caused by entering molecules. The kinetic parameters of substrate entry and exit are derived from a two-state Markov model, analyzing the substrate concentration dependence of the average ion current and its variance. Including filter corrections allows one to detect the transition rates beyond the cutoff frequency, fc, of the instrumental ion-current filter. As an application of the method, we performed an analysis of the single-channel ion current of Meropenem, an antibiotic of the carbapenem family, interacting with OmpF, the major general outer membrane channel of Escherichia coli bacteria. At 40 °C we detected the residence time of Meropenem inside OmpF of about 500 ns - more than 2 orders of magnitude smaller than fc-1 and close to the diffusion limit of few hundred nanoseconds. We also have established theoretical limit conditions under which the substrate-induced channel blockages can be detected and suggest that submicrosecond-scale gating kinetic parameters are accessible with existing experimental equipment

Macroscopic electric field inside water-filled biological nanopores

Multi-drug resistance bacteria are a challenging problem of contemporary medicine. This is particularly critical for Gram-negative bacteria, where antibiotics are hindered by the outer membrane to reach internal targets. Here more polar antibiotics make use of nanometric water-filled channels to permeate inside. We present in this work a computational all-atom approach, using water as a probe, for the calculation of the macroscopic electric field inside water-filled channels. The method allows one to compare not only different systems but also the same system under different conditions, such as pH and ion concentration. This provides a detailed picture of electrostatics in biological nanopores shedding more light on how the charged residues of proteins determine the electric field inside, and also how medium can tune it. These details are central to unveil the filtering mechanism behind the permeation of small polar molecules through nanometric water-filled channel

New Perspectives for Neutron Capture Radiation Therapy with 7Be. The Chemistry and Biochemistry Gap

The research on new radiopharmaceuticals for therapy of cancer is evolving rapidly. Thanks to novel technologies and new selective and less toxic compounds, we move towards personalized molecular medicine. The neutron capture radiation therapy (NCT) can be potentially much safer and can offer a better spatial and temporal control than the radioisotope therapy. Still, there are not many options in NCT: the 10B isotope has been almost exclusively used for decades, and only recently, 157Gd has attracted some interest. Here, we want to draw attention to a new nuclide, 7Be, recently suggested for the NCT, and discuss perspective of Be2+ confinement in aqueous solutions and targeted delivery to cancerous tissues

Glucose transport via the pseudomonad porin OprB: implications for the design of Trojan Horse anti-infectives

Deciphering the transport through outer-membrane porins is crucial to understand how anti-infectives enter Gram-negative bacteria and perform their function. Here we elucidated the transport mechanism of substrates through the Pseudomonads sugar-specific porin OprB by means of multiscale modeling. We used molecular dynamics simulations to quantify the energetics of transport and thus a diffusion model to quantify the macroscopic flux of molecules through OprB. Our results show that Trp171 and several glutamate residues in the constriction region are key for the transport of glucose, the preferred natural substrate, through OprB. The unveiled transport mechanism suggests that 2-acetamido-1,2-dideoxynojirimycin (DNJ-NAc), an anti-infective structurally similar to glucose, can enter the cell via OprB. We quantified its energetics and macroscopic flux through OprB providing a comparative analysis with the natural substrate. Thus this pore can be considered as a promising gateway for exploiting the Trojan Horse strategy in pathogenic bacteria

Exploiting the porin pathway for polar compound delivery into Gram-negative bacteria

Background: In Gram-negative bacteria, the outer-membrane represents an additional barrier for antibiotics to permeate inside pathogens. Our inability to come up with novel effective antibiotics mostly relies upon insufficient understanding of the molecular basis behind outer-membrane penetration. Results: Polar antibiotics can permeate through water-filled porins, such as OmpF and OmpC from Escherichia coli. Through molecular modeling, permeation of imipenem and meropenem was found to be strongly dependent upon capability of drugs to properly align their electric dipole to the internal electric field in the restricted region of the pore. Electrostatics differences between OmpF and OmpC, and modifications along a series of OmpC mutants from E. coli-resistant clinical strains identify a 'preorientation' region, which dramatically affects antibiotic pathway. Conclusion: A novel perspective is presented, suggesting new molecular properties to be included in drug design