BPS2026 – Understanding biofuel-related compounds-induced membrane stress in Zymomonas mobilis at the molecular level

The industrial production of biofuels from lignocellulosic biomass is hindered by inhibitory compounds generated during biomass pretreatment. Zymomonas mobilis , a promising ethanologenic bacterium with high ethanol productivity, is sensitive to inhibitors from several classes of small molecules, including organic acids, aldehydes, alcohols, ketones, and amides. These compounds interact with the bacterial membrane, altering its biophysical properties and compromising cellular function. While experimental studies have provided valuable insights into stress responses in Z. mobilis , the molecular-level mechanisms by which these compounds affect membrane integrity remain unclear. In this study, we employed all-atom molecular dynamics (MD) simulations to investigate interactions of small molecules with a biologically relevant Z. mobilis membrane model. Classical MD simulations were first performed for eight compounds representing carboxylic acids, alcohols, and aldehydes across a concentration range of 0–2.5 mol%. Analysis of key membrane properties, including area per lipid, membrane thickness, lipid order parameter (S CH ), and lateral diffusion coefficient (D xy ), showed that all tested inhibitors altered membrane dynamics, with effects dependent on molecular structure. To expand these findings, we performed replica exchange umbrella sampling (REUS) simulations on thirty-three lignocellulosic inhibitors to calculate free-energy profiles, diffusivities, and membrane permeabilities. Hydrophobicity emerged as the dominant factor governing permeability, and mapping these values against experimentally determined inhibitor concentrations revealed the transport landscape of these compounds. Together, these results provide a molecular framework for understanding how lignocellulosic inhibitors interact with the Z. mobilis membrane. This knowledge offers useful guidance for engineering more tolerant strains and for improving the efficiency of biofuel production processes.