Harnessing Yeast Peroxisomes for Biosynthesis of Fatty-Acid-Derived Biofuels and Chemicals with Relieved Side-Pathway Competition

Abstract

Establishing efficient synthetic pathways for microbial production of biochemicals is often hampered by competing pathways and/or insufficient precursor supply. Compartmentalization in cellular organelles can isolate synthetic pathways from competing pathways, and provide a compact and suitable environment for biosynthesis. Peroxisomes are cellular organelles where fatty acids are degraded, a process that is inhibited under typical fermentation conditions making them an interesting workhouse for production of fatty-acid-derived molecules. Here, we show that targeting synthetic pathways to peroxisomes can increase the production of fatty-acid-derived fatty alcohols, alkanes and olefins up to 700%. In addition, we demonstrate that biosynthesis of these chemicals in the peroxisomes results in significantly decreased accumulation of byproducts formed by competing enzymes. We further demonstrate that production can be enhanced up to 3-fold by increasing the peroxisome population. The strategies described here could be used for production of other chemicals, especially acyl-CoA-derived molecules.

Peroxisomal Dysfunction in Age-Related Diseases

“…Furthermore, additional work is needed to determine whether an increased peroxisomal compartmentalization of metabolic pathways is a mechanism exploited by cancer cells to improve the efficiency of biomass production. Lending support to this notion, compartmentalization of heterologous metabolic pathways in yeast peroxisomes dramatically improves product titer, presumably due to isolation of pathways in a compact and suitable environment [87,88]. …”

Functional screening of aldehyde decarbonylases for long-chain alkane production by Saccharomyces cerevisiae

“…In engineered microbial strains, expression of ADs from a plant (Arabidopsis CER1), an insect ( Drosophila melanogaster CYP4G1), and various species of cyanobacteria (ADOs) displayed long-chain alkane products [12, 13, 16, 17]. However, the low enzyme activities of cyanobacteria ADs have been noticed and only allow for low alkane titers in S. cerevisiae [17–20]. To date, no direct comparative study of ADs from different origins for alkane biosynthesis has been carried out, so we performed a functional screening of different ADs to identify applicable enzyme candidates that can increase alkane production in S. cerevisiae .…”

Enhancement of malate production through engineering of the periplasmic rTCA pathway in Escherichia coli

“…Compartmentalization engineering is a direct approach to limit cross‐talk between the engineered pathways and the cellular milieu (Chen et al, ). Organelles can be used for compartmentalization, such as the mitochondria (Avalos et al, ), chloroplasts (Kumar et al, ), vacuoles (Bayer et al, ), and peroxisomes (DeLoache et al, ; Zhou et al, ). Therefore, we speculated that the periplasm could be used for compartmentalization, as it is isolated from the cytosol.…”

“…Recently, compartmentalization engineering has been adopted at the molecular level to configure and control enzymes in the substrate transmission channel in organelles in eukaryotic cells. Organelles such as mitochondria (Avalos, Fink, & Stephanopoulos, 2013), chloroplasts (Kumar et al, 2012), vacuoles (Bayer et al, 2009), peroxisomes (DeLoache, Russ, & Dueber, 2016;Zhou et al, 2016), and the endoplasmic reticulum (Murakami et al, 2015), which can be isolated from the cytosol with specialized metabolic reactions, can be modified or mimicked to improve engineered pathways (Chen et al, 2018). Unfortunately, these organelles are not present in E. coli (Madigan, 2012), thus leading to some obstacles in applying compartmentalization strategies.…”