Anaerobic fermentation of glycerol by Escherichia coli: a new platform for metabolic engineering

The worldwide surplus of glycerol generated as inevitable byproduct of biodiesel fuel and oleochemical production is resulting in the shutdown of traditional glycerol-producing/refining plants and new applications are needed for this now abundant carbon source. In this article we report our finding that Escherichia coli can ferment glycerol in a pH-dependent manner. We hypothesize that glycerol fermentation is linked to the availability of CO(2), which under acidic conditions is produced by the oxidation of formate by the enzyme formate hydrogen lyase (FHL). In agreement with this hypothesis, glycerol fermentation was severely impaired by blocking the activity of FHL. We demonstrated that, unlike CO(2), hydrogen (the other product of FHL-mediated formate oxidation) had a negative impact on cell growth and glycerol fermentation. In addition, supplementation of the medium with CO(2) partially restored the ability of an FHL-deficient strain to ferment glycerol. High pH resulted in low CO(2) generation (low activity of FHL) and availability (most CO(2) is converted to bicarbonate), and consequently very inefficient fermentation of glycerol. Most of the fermented glycerol was recovered in the reduced compounds ethanol and succinate (93% of the product mixture), which reflects the highly reduced state of glycerol and confirms the fermentative nature of this process. Since glycerol is a cheap, abundant, and highly reduced carbon source, our findings should enable the development of an E. coli-based platform for the anaerobic production of reduced chemicals from glycerol at yields higher than those obtained from common sugars, such as glucose.