Metabolic Flux Analysis for the Biosynthesis of Fuel Hydrocarbons

Timespan: 
2009-2011
Principal Investigator: 

We propose to develop the direct production of fuel hydrocarbons from cellulose-based waste feedstock using a carbon neutral fermentation process employing Gliocladium roseum (NRRL 50072), an endophytic fungus recently isolated from Northern Patagonia. The fungus produces and excretes an extensive series of straight chained and branched medium chain-length hydrocarbons. It can produce these products when grown on cellulose — the world’s most abundant organic compound produced from CO2 fixation by plants. The biofuel produced by isolate NRRL 50072 is not yeast ethanol, but instead a hydrocarbon collection with superior properties for use as a liquid based fuel. Our long-term goal is to characterize the biosynthesis of these products and optimize their production. To achieve this goal it is necessary to develop a comprehensive view of the metabolite biosynthesis pathways of NRRL 50072, including the pathways for substrate to product conversion and the on-path and off-path intermediates produced during fermentation. In this proposal we will outline how the metabolome will be reconstructed through the combined use of liquid chromatography-mass spectroscopy (LC/MS/MS) and nuclear magnetic resonance (NMR) spectroscopic approaches.

This work will be done in a new multidisciplinary collaboration between Professors Scott Strobel in the Department of Molecular Biophysics and Biochemistry and Gary Cline in the Department of Internal Medicine. The proposed research will foster collaboration between two laboratories with interests in the control of metabolism for specialized cellular functions. The Strobel lab has initiated a program to explore NRRL 50072 gas production that includes genomic sequencing, transcriptome analysis, volatile organic compound (VOC) analysis, culturing, and metabolic labeling experiments. Strobel has established collaborations with engineering and bioinformatics groups, which are supported through other funding mechanisms. The Cline lab has traditionally focused on diabetes and has developed methodologies used to identify metabolic pathways that couple metabolism with insulin secretion in pancreatic islet cell lines. These techniques developed by the Cline lab are well-suited to address the metabolic questions of NRRL 50072 biofuel production. The application of these methodologies to alternative energy questions is a new research direction for the Cline lab and a new approach for hydrocarbon analysis for the Strobel lab.

The “mycodiesel” alternative energy technology outlined in this proposal is currently being investigated exclusively at Montana State University and Yale University. The target organism was discovered at MSU and there is a cooperative license and technology agreement in place between the two universities. Consequently, isolate NRRL 50072 exists in cultured form solely at MSU and Yale. Although there are several technologies that seek to obtain biofuels from lignocellulose, the majority are focused upon production of ethanol. A technology capable of producing the medium chain hydrocarbons (C6-C16) present within fossil fuels would be an original and highly valuable technology when successfully developed.

Scott Strobel, Molecular Biophysics and Biochemistry
Gary Cline, Internal Medicine, Division of Endocrinology Metabolism, YNHH