Endocytosing Escherichia coli as a Whole-Cell Biocatalyst of Fatty Acids

Abstract

Whole cell biocatalysts can be used to convert fatty acids into various value-added products. However, fatty acid transport across cellular membranes into the cytosol of microbial cells limits substrate availability and impairs membrane integrity, which in turn decreases cell viability and bioconversion activity. Because these problems are associated with the mechanism of fatty acid transport through membranes, a wholecell biocatalyst that can form caveolae-like structures was generated to promote substrate endocytosis. Caveolin-1 (CAV1) expression in Escherichia coli increased both the fatty acid transport rate and intracellular fatty acid concentrations via endocytosis of the supplemented substrate. Furthermore, fatty-acid endocytosis alleviated substrate cytotoxicity in E. coli. These traits attributed to bacterial endocytosis resulted in dramatically elevated biotransformation efficiencies in fed-batch and cellrecycle reaction systems when caveolae-forming E. coli was used for the bioconversion of ricinoleic acid (12-hydroxyoctadec-9-enoic acid) to (Z)-11-(heptanoyloxy) undec-9-enoic acid. We propose that CAV1-mediated endocytosing E. coli represents a versatile tool for the biotransformation of hydrophobic substrates.

Microbial fatty acid transport proteins and their biotechnological potential

“…Finally, in another study, FadL was overexpressed in a strain expressing human Cav1 proteins (Shin et al, 2019). Cav1 proteins stimulate the formation of endosomes that excise from the inner membrane.…”

“…This study also observed negative effects on overall productivity after a stronger overexpression of FadL, further demonstrating the importance of fine‐tuning the expression of transporters to avoid membrane stress. Finally, in another study, FadL was overexpressed in a strain expressing human Cav1 proteins (Shin et al, 2019). Cav1 proteins stimulate the formation of endosomes that excise from the inner membrane.…”

Production of Rainbow Colorants by Metabolically Engineered Escherichia coli

“…Cav1 has also been employed to increase the accumulation of membrane proteins in E. coli [ 21 ] or to enhance the uptake and conversion of fatty acids into esters. [ 22 ] We thus examined whether the production of rainbow colorants can be enhanced by expanding membrane structures by generation of IMVs through employing caveolae.…”

“…Several recent studies attempted to resolve this issue by membrane engineering [17] such as increasing the membrane area [18,19] or by forming intracellular lipid droplets which can serve as an intracellular reservoir for hydrophobic compounds. [20] Although there have been preliminary reports on employing morphology engineering and formation of inner-membrane vesicles (IMVs) or outer-membrane vesicles (OMVs) for enhancing the production of hydrophobic compounds, [18][19][20][21][22][23] combinatorial applications of these membrane engineering strategies have not been reported. Despite these attempts, the titers of natural colorants need to be further increased for industrial applications.…”

Reprogramming Escherichia coli Metabolism for Bioplastics Synthesis from Waste Cooking Oil

“…Cytotoxicity has been closely associated with reduced metabolic activity, due to substrate or product inhibition. To overcome this issue, several strategies have been developed, such as adding adsorbent resin, 16 forming caveolae, 32 and modifying the membrane structure. Instead, this study employed laboratory adaptive evolution to improve cell tolerance toward elevated FAs concentrations.…”