SYNBIOCHEM

The overall aim of this project is use a synthetic biology approach to develop a consolidated bioprocess (CBP) that will be built around a novel bacterial strain, possessing both high cellulolytic potency and the ability to produce valuable chemicals at high yield. To our knowledge, a fully bi-functional microorganism of this type, able to degrade pretreated cellulosic feedstocks and create useful products (other than ethanol), has so far never been made. Therefore, SYNBIOCHEM proposes to drive biorefining towards hitherto unreached targets. It is now widely recognized that the future bio-economy will rely on cellulosic feedstocks, and that these feedstocks will necessarily be drawn from the abundant non-food, lignocellulosic plant matter (or LC biomass) or from wastes coming from agriculture or recycled papers and cardboards. The use of these cellulosic feedstocks presents a number of challenges for today’s panoply of technologies. These include the initial cracking (pretreatment) of the feedstocks and also the enzymatic hydrolysis of the resultant cellulose, both of which are operations that weigh heavily in the cost balance. One route towards reducing the cost of cellulosic feedstocks processing is to aim for maximum integration, eliminating enzyme production costs, enzyme end-product inhibition and separate hydrolysis and fermentation steps. These elements are all present in a concept known as consolidated bioprocessing (or CBP), which employs a single cellulolytic microorganism as the catalyst for both hydrolysis, leading to the production of glucose from cellulose, and fermentation, leading to the production of target compounds. So far, all the natural cellulolytic organisms produced only limited number of chemicals (namely ethanol, lactate, butyrate and acetate) while the microorganisms engineered to produce higher value chemicals (like Escherichia coli or Saccharomyces cerevisiae) are naturally non-cellulolytic. Furthermore, the production at high yield of some oxidized chemicals derived from acetyl-coA would require the use of a growth associated non-oxidative glycolytic pathway (NOG) under anaerobic conditions but so far all the attempts to develop such a strain have been unsuccessful. In SYNBIOCHEM Clostridum acetobutylicum will form the basis for a sophisticated synthetic biology approach. This bacterium is a very suitable candidate for CBP, owing to its previous industrial history (used in the ABE process), its ability to produce useful chemicals and the presence in its genome of the vital elements necessary to construct a cellulosome. To achieve this goal, we will first repair the cellulosome of C. acetobutylicum and maximize the expression of its encoding genes to degrade efficiently pretreated cellulosic feedstocks and provide fermentable sugars for the production of targeted chemicals. In addition to the creation of a CBP microorganism, SYNBIOCHEM will also provide new knowledge to implement a growth associated and still functional synthetic NOG pathway that will allow the production acetyl-coA derived molecules at higher yield when less NADH is consumed in their formation than produced in the glycolytic pathway. Furthermore, C. acetobutylicum will be metabolically engineered to produce the targeted molecules of industrial interest using as a platform strain, an hydrogenase minus strain previously patented by the applicant. Finally a novel CBP bioreactor will be developed in collaboration with the partners companies. To achieve all of the above targets, SYNBIOCHEM will use a powerful set of synthetic biology tools that have been previously developed during the last four years. Furthermore, in term of knowledge transmission, the professors involved in SYNBIOCHEM will share the expertise gained from this project with the students enrolled in the Master of Biochemical Engineering at INSA and the future International Master of Industrial Biotechnology in collaboration with AgroParisTech.