Fig. 2
Increased expression of fadL increases membrane integrity, fatty acid tolerance and production. a Increased expression of fadL from a plasmid (Pla-fadL) or a genomic insertion (Gen-fadL) both increase the specific growth rate relative to the corresponding controls (Pla-empty, Gen-empty) during challenge with 10 mM C8. Inset values are the specific growth rate, h−1. b Percentage of cells with intact membrane (membrane integrity), assessed using SYTOX Green. Strains with increased expression of fadL, Pla-fadL and Gen-fadL, have improved membrane integrity relative to their corresponding controls, Pla-empty and Gen-empty, during challenge with 10 mM C8. c Strains with increased expression of fadL, Pla-fadL and Gen-fadL, produce more fatty acid than the corresponding controls, Pla-empty and Gen-empty. d The fadL mRNA relative abundance at 48 h has a positive relationship with the fatty acids titer after 72 h. Four different promoters (M1-12, M1-37, M1-46 and M1-93) were used to replace the native promoter of fadL. The mRNA abundance of fadL in M1-12-fadL strain was set as 1. The 16S rrsA gene was used as normalizing factor. For a and b, experiments were performed in shake flasks containing MOPS + 2% (wt/v) dextrose with 10 mM octanoic acid (C8) at an initial pH of 7.0, shaken at 220 rpm, and maintained at 30 °C. For c and d, 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; Pla-fadL: MG1655 + pACYC184-Kan-fadL; Gen-empty: MG1655 ldhA::FRT-cat-FRT; Gen-fadL: MG1655 ldhA::FRT-cat-FRT, fadL. TE: pXZ18Z plasmid