This Small Business Technology Transfer (STTR) Phase I project will develop a facultative anaerobic bacterium that ferments sugars obtained from cellulosic sources into isoprene, a valuable feedstock used in the production of latex rubber, plastics, and pharmaceuticals. Isoprene also is used for the production of chemicals, or polymerized into replacements for petroleum-derived jet fuel. This work will build on prototype E. coli work by redesigning gene constructs and expressing them in the facultative anaerobic host Bacillus coagulans. Phase I of this research will entail synthesizing and cloning a set of isoprene synthase (IspS) genes into B. coagulans; this work will first evaluate ten separate potential IPS molecules to determine the best one for isoprene production in bacteria. The engineered B. coagulans strains will be evaluated for isoprene production by GC-MS headspace analysis. Achieving anaerobic isoprene production from lignocellulosic sources within at least an order of magnitude of the rates achieved in E. coli at the end of Phase I will demonstrate feasibility. Phase II will then focus on improving the flux of metabolites through the pathway to this enzyme. The broader impact/commercial potential of this project will be to develop a high efficiency bacterial strain of Bacillus coagulans to ferment sugars derived from cellulosic materials, such as pulp and pulp mill sludge into isoprene. The paper industry produces an estimated 2 million tons of fiber sludge annually. This material is either discarded in a landfill or burned, adding millions of dollars to the cost of paper production. The composition of this sludge is ~50% cellulose and hemicellulose on a dry weight basis, making it an excellent fermentation feedstock when converted into its constituent five and six carbon sugars. Isoprene is a platform chemical that is capable of playing a central role in the future bio-economy. Annual global consumption of isoprene is 1.7 billion pounds per year, of which more than 95% is used in the production of isoprene rubber, styrene-isoprene-styrene block copolymer (SIS), and butyl rubber, with an estimated price of $1.30 per pound. The balance is used in the production of high-value isoprenoid derivatives such as vitamins, nutraceuticals, pesticides, fragrances, flavors and pharmaceuticals. The project also will provide an improved understanding of the biosynthetic pathway of isoprene and other high-value isoprenoid compounds.
