Improving Microbial Biogasoline Production in Escherichia coli Using Tolerance Engineering

Abstract

Engineering microbial hosts for the production of fungible fuels requires mitigation of limitations posed on the production capacity. One such limitation arises from the inherent toxicity of solvent-like biofuel compounds to production strains, such as Escherichia coli. Here we show the importance of host engineering for the production of short-chain alcohols by studying the overexpression of genes upregulated in response to exogenous isopentenol. Using systems biology data, we selected 40 genes that were upregulated following isopentenol exposure and subsequently overexpressed them in E. coli. Overexpression of several of these candidates improved tolerance to exogenously added isopentenol. Genes conferring isopentenol tolerance phenotypes belonged to diverse functional groups, such as oxidative stress response (soxS, fpr, and nrdH), general stress response (metR, yqhD, and gidB), heat shock-related response (ibpA), and transport (mdlB). To determine if these genes could also improve isopentenol production, we coexpressed the tolerance-enhancing genes individually with an isopentenol production pathway. Our data show that expression of 6 of the 8 candidates improved the production of isopentenol in E. coli, with the methionine biosynthesis regulator MetR improving the titer for isopentenol production by 55%. Additionally, expression of MdlB, an ABC transporter, facilitated a 12% improvement in isopentenol production. To our knowledge, MdlB is the first example of a transporter that can be used to improve production of a short-chain alcohol and provides a valuable new avenue for host engineering in biogasoline production.

RNA modification enzymes encoded by the gid operon: Implications in biology and virulence of bacteria

“…Interestingly, a recent study has implicated GidB in the improvement of microbial biogasoline production [122]. Isopentenol is an important target compound in pharmaceutical production, and is also a useable biofuel [123].…”

“…The biggest challenge to microbial production of such compounds is the large amount of biomass needed to make large quantities, and isopentenol production in E. coli has shown to be relatively low [124,125]. The study by Foo et al identified 40 genes in E. coli that were up-regulated after exposure to isopentenol [122]. Of the 40 genes, gidB was one of six genes that increased both isopentenol tolerance and production in E. coli.…”

Integrated analysis of isopentenyl pyrophosphate (IPP) toxicity in isoprenoid-producing Escherichia coli

“…Considerable progress has been made in the production of isoprenol, a C5 alcohol derived from isopentenyl pyrophosphate (IPP), which is one of two universal precursors for isoprenoid compounds (George et al, 2015b;Zheng et al, 2013). In E. coli, the toxicity of both isoprenol (Foo et al, 2014) and IPP has been suggested to impact cell growth and limit product yields. Although the toxicity of IPP (and its longer chain derivatives GPP and FPP) was observed over a decade ago (Martin et al, 2003), a systematic assessment of this stress and its impact on host physiology has yet to be conducted.…”

Evolution of a Biomass-Fermenting Bacterium To Resist Lignin Phenolics

“…Gene expression at a t of 0.5 h shows that abrupt ferulate stress induces expression of genes encoding the efflux pump cphy1055-cphy1056, which is similar to E. coli mdlAB conferring resistance to organic solvents (21). Many of the genes upregulated at a t of 0.5 h are colocated in two genomic regions.…”