Molecular-dynamics-simulation-guided membrane engineering allows the increase of membrane fatty acid chain length in Saccharomyces cerevisiae

Jeroen M Maertens,Samuel Genheden,Lisbeth Olsson,Simone Scrima,Leif A Eriksson,Matteo Lambrughi,Cecilia Trivellin,Maurizio Bettiga,Elena Papaleo

doi:10.1038/s41598-021-96757-y

Jeroen M Maertens, Samuel Genheden + Show 7 more

Open Access

https://doi.org/10.1038/s41598-021-96757-y

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Abstract

The use of lignocellulosic-based fermentation media will be a necessary part of the transition to a circular bio-economy. These media contain many inhibitors to microbial growth, including acetic acid. Under industrially relevant conditions, acetic acid enters the cell predominantly through passive diffusion across the plasma membrane. The lipid composition of the membrane determines the rate of uptake of acetic acid, and thicker, more rigid membranes impede passive diffusion. We hypothesized that the elongation of glycerophospholipid fatty acids would lead to thicker and more rigid membranes, reducing the influx of acetic acid. Molecular dynamics simulations were used to predict the changes in membrane properties. Heterologous expression of Arabidopsis thaliana genes fatty acid elongase 1 (FAE1) and glycerol-3-phosphate acyltransferase 5 (GPAT5) increased the average fatty acid chain length. However, this did not lead to a reduction in the net uptake rate of acetic acid. Despite successful strain engineering, the net uptake rate of acetic acid did not decrease. We suggest that changes in the relative abundance of certain membrane lipid headgroups could mitigate the effect of longer fatty acid chains, resulting in a higher net uptake rate of acetic acid.

Highlights

Abbreviations AOPC 1-Arachidoyl-2-oleoyl-sn-glycero-3-phosphocholine AOPI 1-Arachidoyl-2-oleoyl-sn-glycero-3-phospho‐(1’‐myo‐inositol) area per lipid (APL) Area per lipid BOPC 1-Behenoyl-2-oleoyl-sn-glycero-3-phosphocholine BOPI 1-Behenoyl-2-oleoyl-sn-glycero-3-phospho‐(1’‐myo‐inositol) cytidine diphosphate diacylglycerol (CDP-DAG) Cytidine diphosphate diacylglycerol Cer Ceramide CL Cardiolipin DAG Diacylglycerol DOPC 1,2‐Dioleoyl‐sn‐glycerol‐3‐phosphocholine EE Ergosterol ester ERG Ergosterol fatty acid (FA) Fatty acid glycerol-3-phosphate acyl transferase (GPAT) Glycerol-3-phosphate acyl transferase GPL Glycerophospholipid GV Green value IPC Inositol phosphorylceramide largest part of the molecule (L-GPL) Lyso-glycerophospholipid
Molecular Dynamics (MD) simulations were used to investigate how increasing the concentrations of very-long-chain FA GPLs affect the structural properties of yeast membrane models
This indicates that the FAE1_GPAT5 double expression strain is more susceptible to acetic acid stress compared to the control strain. This could be due to a range of reasons, including, for example, reduced intracellular buffering capacity, or reduced efflux rate. These results, together with shotgun lipidomic data, show complex changes in the engineered strain that we did not include in the MD simulations by which we focus on investigate how increasing concentrations of verylong-chain FA GPLs affect the structural properties of models of yeast membranes

Summary

Introduction

Abbreviations AOPC 1-Arachidoyl-2-oleoyl-sn-glycero-3-phosphocholine AOPI 1-Arachidoyl-2-oleoyl-sn-glycero-3-phospho‐(1’‐myo‐inositol) APL Area per lipid BOPC 1-Behenoyl-2-oleoyl-sn-glycero-3-phosphocholine BOPI 1-Behenoyl-2-oleoyl-sn-glycero-3-phospho‐(1’‐myo‐inositol) CDP-DAG Cytidine diphosphate diacylglycerol Cer Ceramide CL Cardiolipin DAG Diacylglycerol DOPC 1,2‐Dioleoyl‐sn‐glycerol‐3‐phosphocholine EE Ergosterol ester ERG Ergosterol FA Fatty acid GPAT Glycerol-3-phosphate acyl transferase GPL Glycerophospholipid GV Green value IPC Inositol phosphorylceramide L-GPL Lyso-glycerophospholipid. Acetic acid can be very damaging to Saccharomyces cerevisiae and even trigger a poptosis[9,10] It is of the utmost importance to minimize the intracellular acetic acid concentration. In S. cerevisiae, de novo fatty acid (FA) synthesis is catalysed by a complex consisting of Fas[1] and Fas[2] This complex produces acyl chains with lengths of up to eighteen carbon atoms (C18). The glycerophospholipids (GPLs) are the main group of membrane lipids, and their important components are included in Fig. 1B (chemical structures: Additional file 1, S1). Other membrane lipids have characteristic shapes: sphingolipids, phosphatidylcholine (PC), phosphatidylserine (PS) and phosphatidylinositol (PI) are considered cylindrical, while phosphatidic acid (PA), phosphatidylethanolamine (PE) and diacylglycerol (DAG) are considered conical with a small headgroup (Fig. 1A). The shape can affect the membrane structure and, in addition to the different membrane components and their headgroups, FA chain length and degree of saturation are important factors affecting membrane properties such as the area per lipid (APL) and the rigidity of the membrane[18,19]

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