Researchers Engineer Yeast to Produce Methyl Halides from Biomass; Precursors for Biohydrocarbon Fuels

22 April 2009

CH3I production from cellulosic feedstocks using a microbial co-culture. A. fermentans ferments cellulosic feedstocks to acetate and ethanol, which the modified S. cerevisiae uses to produce methyl halides. Adapted from Bayer et al. (2009). Click to enlarge.

Researchers at the University of California San Francisco have engineered the industrial yeast S. cerevisiae to convert biomass to methyl halides with good yield. As end products, methyl halides (CH₃X, X = Cl, Br or I) are used in a variety of applications. They can also be used as intermediates for the chemical synthesis of more complex carbon compounds such as fuel hydrocarbons.

Zeolite catalysts (e.g., ZSM-5 and SAPO-34) have been used to convert methyl halides to products including gasoline, olefins, aromatics, alcohols and ethers. A method to convert biomass to methyl halides thus enables the transformation of biomass into drop-in chemicals and liquid fuels—e.g., bio-gasoline—in a two-step process. A paper on the work was published online 20 April in the Journal of the American Chemical Society.

Methyl halides are naturally produced by a number of organisms; the enzyme responsible for this production—methyl halide transferase (MHT)—has been studied in the context of the environmental production of methyl halides, which contribute to ozone depletion. However, the yield of CH₃X from natural producers is low; moving MHT into an industrial organism is a fundamental requirement for commercialization.

The UCSF team, led by Christopher Voigt, used what they called a “synthetic metagenomic” approach to identify, and synthesize all 89 putative MHT genes from plants, fungi, bacteria and unidentified organisms present in the NCBI sequence database. They then screened this set in Escherichia coli to identify the rates of CH₃Cl, CH₃Br, and CH₃I production, with 56% of the library active on chloride, 85% on bromide, and 69% on iodide.

The MHT from B. maritima displayed the highest activity of all genes. They then transferred the B. maritima MHT to the industrial yeast S. cerevisiae. Testing with glucose and sucrose as a feedstock showed a yield of 190 mg/L^-1 from the modified yeast.

To enable the yeast to use biomass as a feedstock, the team designed a novel co-culture of the MHT-expressing yeast with a mesophyllic cellulolytic bacterium, Actinotalea fermentans, which was isolated from a landfill in France. A. fermentans produces acetate and ethanol.

Acetate and ethanol inhibit the growth of A. fermentans. Thus, the researchers note, they created a metabolic interdependence in the community, with S. cerevisiae dependent on A. fermentans for carbon and energy, and A. fermentans dependent on S. cerevisiae for metabolism of toxic waste products.

Using the symbiotic co-culture of the engineered yeast and A. fermentans, they achieved methyl halide production from unprocessed switchgrass (Panicum virgatum), corn stover, sugar cane bagasse, and poplar (Populus sp).

Construction of a synthetic microbial co-culture, where a cellulolytic bacterium provided acetate to methyl iodide producing yeast, allowed diverse lignocellulosic feedstocks to be converted to methyl halides. This approach differs from other engineered examples of cellulose digestion where single species are engineered to perform cellulose cleavage, followed by utilization of the resulting five- and six-carbon sugars.

The co-culture described here divides the cellulolytic- and methyl halide-producing functions between two species. Metabolic division of labor is commonly observed in microbial consortia, including soil and termite gut microbiota that degrade cellulose, and has been used to engineer strains of Bacillus subtilis that cooperatively produce cellulosomes.

...There are two similar processes that could be used for biomass conversion. The first is the use of microbial methanogens to produce methane, which will then require chemical activation for downstream conversion into liquid fuels. The second is the production of syngas by pyrolysis or microorganisms. Both of these processes require large energy inputs to convert the gas to liquid fuels. We are proposing to bypass the energy-intensive steps of the Fischer-Tropsch process by having cells directly produce activated methane, using the chemical energy in ATP. Methyl halides may prove to be a fungible intermediate through which the carbon from different sources of biomass can be converted into a wide range of chemicals, consumer products, and liquid fuels.

—Bayer et. al. (2009)

Resources

• Travis S. Bayer, Daniel M. Widmaier, Karsten Temme, Ethan A. Mirsky, Daniel V. Santi and Christopher A. Voigt (2009) J. Am. Chem. Soc., Article ASAP doi: 10.1021/ja809461u

• K. R. Redeker, N.-Y. Wang, J. C. Low, A. McMillan, S. C. Tyler, R. J. Cicerone (2000) Emissions of Methyl Halides and Methane from Rice Paddies. Science Vol. 290. no. 5493, pp. 966 - 969 doi: 10.1126/science.290.5493.966