Fig. 3.

Deletion of ompF and increased expression of fadL have an additive effect on increasing membrane integrity, fatty acid tolerance and production. a Combinatorial deletion of ompF (ΔompF) and increased expression of fadL (Pla-fadL) increases the specific growth rate during challenge with 10 mM C8 relative to the starting strain (Pla-empty), individual ompF deletion strain (ΔompF + Pla-empty), and individual overexpression of fadL (Pla-fadL). Inset values are the specific growth rate, h⁻¹ b Percentage of cells with intact membrane (membrane integrity), assessed using SYTOX Green. Combinatorial deletion of ompF and increased expression of fadL improves membrane integrity during challenge with 10 mM C8 relative to Pla-empty, ΔompF + Pla-empty and Pla-fadL strains. c The combined implementation of ompF deletion and increased expression of fadL supports increased fatty acid titers relative to each engineering strategy implemented individually. For a and b, experiments were performed in MOPS + 2% (wt/v) dextrose shake flasks at 220 rpm 30 °C with an initial pH of 7.0, 10 mM octanoic acid (C8). For c, all strains carry the pXZ18Z plasmid (TE, fabZ) for LCFA (C14–C16) production. Fermentations were performed in MOPS + 2% (wt/v) dextrose shake flasks at 220 rpm 30 °C with an initial pH of 7.0, 1.0 mM IPTG. Values are the average of at least three biological replicates with error bars indicating one standard deviation. Percent increase values are shown only for differences that were deemed statistically significant (P < 0.05). Pla-empty: MG1655 + pACYC184-Kan; ΔompF + Pla-empty: MG1655, ΔompF + pACYC184-Kan; Pla-fadL: MG1655 + pACYC184-Kan-fadL; ΔompF + Pla-fadL: MG1655, ΔompF + pACYC184-Kan-fadL