Abstract
Gram-negative bacteria cause the majority of highly drug-resistant bacterial infections. To cross the outer membrane of the complex Gram-negative cell envelope, antibiotics permeate through porins, trimeric channel proteins that enable the exchange of small polar molecules. Mutations in porins contribute to the development of drug-resistant phenotypes. In this work, we show that a single point mutation in the porin PorB from Neisseria meningitidis, the causative agent of bacterial meningitis, can strongly affect the binding and permeation of beta-lactam antibiotics. Using X-ray crystallography, high-resolution electrophysiology, atomistic biomolecular simulation, and liposome swelling experiments, we demonstrate differences in drug binding affinity, ion selectivity and drug permeability of PorB. Our work further reveals distinct interactions between the transversal electric field in the porin eyelet and the zwitterionic drugs, which manifest themselves under applied electric fields in electrophysiology and are altered by the mutation. These observations may apply more broadly to drug-porin interactions in other channels. Our results improve the molecular understanding of porin-based drug-resistance in Gram-negative bacteria.
Highlights
As evidenced by the world-wide Covid19 pandemic, untreatable in fectious diseases have a huge impact on public health, the global economy, and the lives of people across the world
We show that a single point mutation in the porin PorB from Neisseria meningitidis, the causative agent of bacterial meningitis, can strongly affect the binding and permeation of betalactam antibiotics
Most antibiotic drugs permeate the outer membrane of Gramnegative bacteria through porins, beta-barrel proteins that form large water-filled channels in the membrane used by the organisms to import nutrients [7,8,9]
Summary
As evidenced by the world-wide Covid pandemic, untreatable in fectious diseases have a huge impact on public health, the global economy, and the lives of people across the world. We used a combination of X-ray crystallography, BBA - Biomembranes 1863 (2021) 183601 electrophysiology, and molecular dynamics simulations to study the role of mutations in PorB of Neisseria meningitidis (Nme) – the causal agent of bacterial meningitis – on antibiotic binding and permeation. PorB is a trimeric anion-selective porin and the second most abundant protein in the outer membrane of N. meningitidis, possessing a binding site for antibiotics in its pore [14] It plays an important role in the human immune response to neisserial infections and has been linked to inducing apoptosis upon infection of host cells [18,19,20]. The effect of the mutation on reducing ampicillin uptake is shown by liposome-swelling assays
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