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1. Activation of a Bacterial Mechanosensitive Channel, MscL, Underlies the Membrane Permeabilization of Dual-Targeting Antibacterial Compounds

   https://doi.org/10.3390/antibiotics11070970  

   Wray, Robin; Wang, Junmei; Blount, Paul; Iscla, Irene (July 2022, Antibiotics)

   Resistance to antibiotics is a serious and worsening threat to human health worldwide, and there is an urgent need to develop new antibiotics that can avert it. One possible solution is the development of compounds that possess multiple modes of action, requiring at least two mutations to acquire resistance. Compound SCH-79797 both avoids resistance and has two mechanisms of action: one inhibiting the folate pathway, and a second described as “membrane permeabilization”; however, the mechanism by which membranes from bacterial cells, but not the host, are disrupted has remained mysterious. The opening of the bacterial mechanosensitive channel of large conductance, MscL, which ordinarily serves the physiological role of osmotic emergency release valves gated by hypoosmotic shock, has been previously demonstrated to affect bacterial membrane permeabilization. MscL allows the rapid permeabilization of both ions and solutes through the opening of the largest known gated pore, which has a diameter of 30 Å. We found that SCH-79797 and IRS-16, a more potent derivative, directly bind to the MscL channel and produce membrane permeabilization as a result of its activation. These findings suggest that possessing or adding an MscL-activating component to an antibiotic compound could help to lower toxicity and evade antibiotic resistance. 

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2. Fusion peptide from SARS-2 spike transforms into a wedge inserted in a bilayer leaflet, and thins the opposite leaflet

   Steven R Van Doren, Benjamin Scott (February 2022, Biophysical journal)

   Activation of SARS-CoV-2 Spike deploys its fusion peptide to a membrane of the host cell to infect it. NMR in solution demonstrates that this fusion peptide transforms from intrinsic disorder in solution into a wedge-shaped structure inserted in bilayered micelles. According to NOEs and proximity to a nitroxide spin label deep in the membrane mimic, the globular fold of three helices contrasts the open, extended conformations observed in compact prefusion states. In the hydrophobic, narrow end of the wedge, helices 1 and 2 contact the fatty acyl chains of phospholipids. 50 of the resulting paramagnetic NMR relaxation enhancements and 6 lipid-protein NOEs provided ambiguous distances as collective variables (colvars) to bias and guide MD simulations. Simulations in NAMD using the CHARMM36 forcefield included colvars for 130 medium- and long-range NOEs to maintain the equilibrium structure. In the gently NMR-biased simulations, the fusion peptide maintained its insertion of helices 1 and 2 within a single leaflet while helix 3 remained exposed. A cation occasionally visited the anionic side chains in the loop joining helices 2 and 3 or at the N-terminal end of helix 1. The unoccupied leaflet is thinned and distorted opposite the fusion peptide.The thinning could be related to the fusion peptide promoting formation of the hemi-fusion intermediate in the process of viral-cell fusion. Supported by NSF Rapid award 2030473. 

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3. In Silico Screen Identifies a New Family of Agonists for the Bacterial Mechanosensitive Channel MscL

   https://doi.org/10.3390/antibiotics11040433  

   Wray, Robin; Blount, Paul; Wang, Junmei; Iscla, Irene (April 2022, Antibiotics)

   MscL is a highly conserved mechanosensitive channel found in the majority of bacterial species, including pathogens. It functions as a biological emergency release valve, jettisoning solutes from the cytoplasm upon acute hypoosmotic stress. It opens the largest known gated pore and has been heralded as an antibacterial target. Although there are no known endogenous ligands, small compounds have recently been shown to specifically bind to and open the channel, leading to decreased cell growth and viability. Their binding site is at the cytoplasmic/membrane and subunit interfaces of the protein, which has been recently been proposed to play an essential role in channel gating. Here, we have targeted this pocket using in silico screening, resulting in the discovery of a new family of compounds, distinct from other known MscL-specific agonists. Our findings extended the study of this functional region, the progression of MscL as a viable drug target, and demonstrated the power of in silico screening for identifying and improving the design of MscL agonists. 

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4. Rapid constriction of the selectivity filter underlies C-type inactivation in the KcsA potassium channel

   https://doi.org/10.1085/jgp.201812082  

   Li, Jing; Ostmeyer, Jared; Cuello, Luis G.; Perozo, Eduardo; Roux, Benoît (October 2018, The Journal of General Physiology)

   C-type inactivation is a time-dependent process observed in many K + channels whereby prolonged activation by an external stimulus leads to a reduction in ionic conduction. While C-type inactivation is thought to be a result of a constriction of the selectivity filter, the local dynamics of the process remain elusive. Here, we use molecular dynamics (MD) simulations of the KcsA channel to elucidate the nature of kinetically delayed activation/inactivation gating coupling. Microsecond-scale MD simulations based on the truncated form of the KcsA channel (C-terminal domain deleted) provide a first glimpse of the onset of C-type inactivation. We observe over multiple trajectories that the selectivity filter consistently undergoes a spontaneous and rapid (within 1–2 µs) transition to a constricted conformation when the intracellular activation gate is fully open, but remains in the conductive conformation when the activation gate is closed or partially open. Multidimensional umbrella sampling potential of mean force calculations and nonequilibrium voltage-driven simulations further confirm these observations. Electrophysiological measurements show that the truncated form of the KcsA channel inactivates faster and greater than full-length KcsA, which is consistent with truncated KcsA opening to a greater degree because of the absence of the C-terminal domain restraint. Together, these results imply that the observed kinetics underlying activation/inactivation gating reflect a rapid conductive-to-constricted transition of the selectivity filter that is allosterically controlled by the slow opening of the intracellular gate. 

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5. Molecular Insights into Cholesterol Concentration Effects on Planar and Curved Lipid Bilayers for Liposomal Drug Delivery

   https://doi.org/10.1101/2025.04.06.647491  

   Khodadadi, Ehsaneh; Khodadadi, Ehsan; Chaturvedi, Parth; Moradi, Mahmoud (April 2025, bioRxiv)

   Abstract Liposomal carriers provide a flexible and effective strategy for delivering therapeutics across a broad spectrum of diseases. Cholesterol is frequently included in these systems to improve membrane rigidity and limit permeability. Despite its widespread use, the optimal cholesterol-to-lipid proportion for achieving stable and efficient liposome performance remains to be fully determined. In this work, we apply all-atom molecular dynamics simulations to explore how different cholesterol concentrations influence the structural and dynamic characteristics of 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) bilayers, considering both planar and curved membrane geometries. Bilayers with cholesterol molar ratios of 0%, 10%, 20%, 30%, 40%, and 50% were simulated, and key biophysical parameters including area per lipid (APL), membrane thickness, leaflet interdigitation, and deuterium order parameters (SCD) were analyzed. In planar bilayers, increasing cholesterol concentration led to a progressive decrease in APL from approximately 60^°A²to 40^°A², accompanied by increased membrane thickness and lipid ordering, consistent with cholesterol’s classical condensing effect. In contrast, curved bilayers exhibited a cholesterol-induced expansion effect, particularly in the inner leaflet, where APL increased from approximately 60^°A²to 90^°A²with rising cholesterol levels. SCD profiles showed that cholesterol enhanced tail ordering up to 40% concentration, beyond which the effect plateaued or slightly declined, suggesting structural saturation or packing frustration. Membrane thickness displayed a monotonic increase in planar bilayers but followed a nonlinear trend in curved systems due to curvature-induced stress. These findings highlight that cholesterol’s influence on membrane properties is highly dependent on bilayer geometry and asymmetry. While planar bilayers exhibit predictable responses, curved systems reveal nonclassical behaviors that challenge traditional models of cholesterol-lipid interactions. This work provides molecular-level insights and establishes a computational framework for the rational design of liposomal systems, emphasizing the need to account for curvature and asymmetry in membrane engineering. 

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