# Quantifying Membrane Structure and Dynamics during Bioproduct Production in Zymomonas mobilis by Molecular Simulation

## TL;DR

This study uses molecular simulations to explore how small molecules from biomass affect the membrane of Zymomonas mobilis, a bacterium used in biofuel production.

## Contribution

The study provides a mechanistic understanding of how small molecules disrupt Zymomonas mobilis membranes, offering insights for strain engineering.

## Key findings

- Membrane thickness decreases and area per lipid increases with higher concentrations of small molecules.
- Hydrophobic molecules like isobutanol cause stronger membrane perturbations.
- Hopanoids stabilize membranes for small molecules but are ineffective against larger hydrophobic ones.

## Abstract

The conversion of
lignocellulosic biomass into biofuels
and bioproducts
by microbial biorefineries is central to a sustainable chemical industry. Zymomonas mobilis is one such biorefinery chassis
and is resistant to ethanol stress, leading to its use in biomass
conversion to biofuels and bioproducts. However, Z.
mobilis growth is often inhibited by organic acids,
aldehydes, alcohols, ketones, and amides found in biomass hydrolysate.
The resulting slow growth inhibits production and as a result drives
up the price for the resulting products. One hypothesis is that these
molecules interact with or disrupt the bacterial membrane, triggering
stress responses and hindering growth. To test this hypothesis at
the molecular level, we employ all-atom molecular dynamics (MD) simulations
to investigate lignocellulose-derived small molecules and their impact
on a biologically relevant Z. mobilis membrane model. Simulations were conducted across a range of inhibitor
concentrations from 0 to 2.5 mol %, analyzing key membrane properties
such as area per lipid (APL), membrane thickness, lipid-order parameter
(−S
CH), lateral diffusion coefficient
(D

xy
), and permeability
coefficient (Pm). From simulation, we observed altered membrane structure
and dynamics at these modest small molecule concentrations commonly
found in hydrolysates. Generally, the membranes become thinner, with
a higher area per lipid and lower-order parameter as the small molecule
concentration increases. These trends are stronger for more hydrophobic
molecules with greater hydrophobic bulk, as isobutanol, propanol,
and propanoic acid showed greater membrane perturbations as the concentration
increased compared to other small molecules. Tracking small molecule
distributions directly in our equilibrium simulations allows us to
determine concentration-dependent free energy profiles for these molecules.
While the trends are noisy, generally the barriers to crossing the
membrane decrease as the concentration increases, indicating that
the membranes become leakier as small molecule concentrations rise.
Comparing between native Z. mobilis membranes with hopanoids and membranes sharing the same phospholipid
composition but without hopanoids, hopanoids stabilize and order the
membrane for smaller molecules to maintain membrane structure but
appear insufficient for larger hydrophobic molecules like isobutanol.
These findings provide a mechanistic understanding of how small molecules
found in biomass degradation streams interact with the Z. mobilis membrane, offering valuable insights for
future strain engineering efforts to optimize biofuel and bioproduct
synthesis from biomass feedstocks by highlighting limits to small
molecule tolerance. This knowledge can guide the modification of membrane
composition to develop more robust microbes, thereby improving microbial
survival and yields in industrial contexts.