| Introduction: The need to reduce CO2 emissions has promoted the use of renewable resources as starting material for the production of fuels and chemicals. The acetone-butanol-ethanol fermentation (ABE fermentation), which has been once industrial, has a great potential for the production of butanol from lignocellulosic biomass. Butanol represents an important source of additives to be blended with existing fuels, and therefore contributes to meeting the increasing demand for renewable fuels (figure 1). Next to that it is also a widely used solvent and chemical feedstock. However, to again make the ABE process economically viable there are some improvements to be achieved; higher butanol yields and higher volumetric productivity to reduce recovery costs. Aim: We want, by means of genetic engineering of existing acetone-butanol-ethanol-producing Clostridial species, to increase the butanol yield and reduce by product formation of acids and acetone. Research: Genetic tools are being developed to engineer a Clostridium acetobutylicum strain with a single-route fermentation pathway. This single-route mutant should convert glucose into 1 butanol, with a theoretical maximum yield of 1 mol BuOH/mol glucose. The wild type metabolic network includes enzymes that catalyse competing reactions whose production is at the expense of butanol production. These products include acids such as lactate, acetate, and butyrate, but also solvents like ethanol and acetone. Enzymes involved in these reactions are our first knock out targets. Generated (multiple) knock out mutants of C. acetobutylicum will be characterized in batch fermentations (laboratory scale 1 2 L bioreactors), by functional genomics (DNA Microarray analysis) as well as by routine physiological analyses. Future research: First our gene knock out system is being optimised and validated. This will give us the ability to generate mutants with multiple knocked out genes. Metabolic modelling of these mutants should provide us with information on further optimisation of the metabolic pathways involved in butanol formation. This project is financially supported by the Netherlands Ministry of Economic Affairs and the B-Basic partner organizations (www.b-basic.nl) through B-Basic, a public-private NWO-ACTS programme (ACTS = Advanced Chemical Technologies for Sustainability). |
