Production of biorenewable styrene: utilization of biomass-derived sugars and insights into toxicity

Abstract

Fermentative production of styrene from glucose has been previously demonstrated in Escherichia coli. Here, we demonstrate the production of styrene from the sugars derived from lignocellulosic biomass depolymerized by fast pyrolysis. A previously engineered styrene-producing strain was further engineered for utilization of the anhydrosugar levoglucosan via expression of levoglucosan kinase. The resulting strain produced 240 ± 3 mg L(-1) styrene from pure levoglucosan, similar to the 251 ± 3 mg L(-1) produced from glucose. When provided at a concentration of 5 g L(-1), pyrolytic sugars supported styrene production at titers similar to those from pure sugars, demonstrating the feasibility of producing this important industrial chemical from biomass-derived sugars. However, the toxicity of contaminant compounds in the biomass-derived sugars and styrene itself limit further gains in production. Styrene toxicity is generally believed to be due to membrane damage. Contrary to this prevailing wisdom, our quantitative assessment during challenge with up to 200 mg L(-1) of exogenously provided styrene showed little change in membrane integrity; membrane disruption was observed only during styrene production. Membrane fluidity was also quantified during styrene production, but no changes were observed relative to the non-producing control strain. This observation that styrene production is much more damaging to the membrane integrity than challenge with exogenously supplied styrene provides insight into the mechanism of styrene toxicity and emphasizes the importance of verifying proposed toxicity mechanisms during production instead of relying upon results obtained during exogenous challenge.

Studies on styrene concentration in drinking water and hot beverages in some settings

“…The world's plastic production is consequentially increasing every year (Jambeck et al, 2015). The total global consumption of styrene is approximately 25 million metric tons annually, which estimates about 30 billion in the USD market (Lian et al, 2016). About 1.5 million tons of plastic is produced yearly by the bottled water industry alone.…”

Emerging tools, enabling technologies, and future opportunities for the bioproduction of aromatic chemicals

“…Progress is already being made to this end. Styrene production, for example, has been demonstrated directly from lignocellulosic biomass depolymerized via fast pyrolysis . Specifically, a previously engineered styrene‐producing E. coli strain was further engineered to co‐expression of levoglucosan kinase (encoded by lgk from Lipomyces starkeyi ), enabling utilization of levoglucosan, an anhydrosugar and the major pyrolytic sugar derived from red oak.…”

“…[96][97][98][99][100] Production of styrene in E. coli, for example, was found to induce membrane damage and leakage (contrastingly, exogenous addition of styrene triggered little change in membrane integrity). 101 In addition to this common and general mechanism, inhibition imposed by aromatics can also further depend upon structural and functional group diversity. For instance, in addition to damaging cell membranes, p-coumaric acid is reported to also interact with DNA, negatively affecting replication and transcription.…”

Engineering membrane and cell-wall programs for tolerance to toxic chemicals: Beyond solo genes

“…Damage to the cell membrane has long been implicated in the toxicity of various chemicals, with the degree of toxicity correlating to the hydrophobicity[12,13]. Short alcohols and hydrophobic compounds, such as ethanol or butanol, can partition into lipid bilayer membranes[14,15]. These compounds displace water at the membrane/water interface which reduces surface tension.…”