New JBEI Methodology Speeds Search for Cellulosic Biofuel Microbes

20 March 2009

A new analytical technique developed by researchers at the US Department of Energy (DOE) Joint BioEnergy Institute (JBEI) promises to speed up greatly the search for microbes that can ferment complex cellulosic sugars under the harsh conditions of biofuels production, such as high temperature, and do not become inhibited by the fuel being produced.

One potential candidate—Geobacillus thermoglucosidasius—has already emerged and JBEI researchers have made important determinations about its metabolism via the novel experimental route.

Our results show that the recently discovered extremophile—the bacterium Geobacillus thermoglucosidasius—which thrives in the high temperatures and pressures of petroleum reservoirs, can ferment the major C₅ and C₆ sugars (e.g., xylose and glucose) in cellulosic biomass and can tolerate high concentrations of ethanol. These capabilities make Geobacillus thermoglucosidasius an ideal microbe for improved production of bio-ethanol and other more advanced biofuels.

—Dr. Jay Keasling, CEO of JBEI

Working with Keasling on this study were Rajat Sapra, a biochemist with Sandia National Laboratories who directs the enzyme optimization program at JBEI, and Yinjie Tang, a chemical engineer now with Washington University in St. Louis. Their results were reported in a paper in the 1 April 2009 issue of the journal Biotechnology and Bioengineering.

Scientists normally obtain metabolic information on a microbe through the sequencing of its genome and subsequent genomic, proteomic and physiological studies—a process that can take months or even years to complete. The new strategy—a combination of in vitro enzyme assays and a unique metabolic flux analysis based on the carbon-13 isotope (¹³C)—significantly shrinks this time frame. The JBEI researchers were able to complete their metabolic studies on G. thermoglucosidasius in less than two weeks.

Metabolic flux is a measurement of the enzymes and reaction activities taking place along a given metabolic pathway as an organism converts its food (such as sugars) into energy and end-products (such as ethanol). The researchers used glucose labeled with carbon-13 to study the distribution of ¹³C in the metabolites formed from glucose. The position of the ¹³C label and the metabolites formed serve as a measurement of metabolic flux in real-time and help determine the metabolic pathways of the organism.

Using our methodology, one can rapidly analyze whether a bacterium is suitable as a biofuels host at an extremely fast time frame as compared to the traditional methods. This is especially important now that we are discovering new and novel bacteria at a pace much, much faster than can be analyzed using the classical techniques. Our isotopomer [isotopic isomer]-based approach for studying cellular metabolisms will not only enable us to identify the best and most promising hosts, but will also provide guidelines for engineering new metabolic pathways that promote biofuel production as well as other environmental and industrial applications.

—Dr. Rajat Sapra

JBEI is one of three US Department of Energy Bioenergy Research Centers. Its scientific partnership is led by Lawrence Berkeley National Laboratory (Berkeley Lab) and includes the Sandia National Laboratories (Sandia), the University of California (UC) campuses of Berkeley and Davis, the Carnegie Institution for Science and the Lawrence Livermore National Laboratory (LLNL).

Resources

• Yinjie J. Tang et al. (2009) Analysis of metabolic pathways and fluxes in a newly discovered thermophilic and ethanol-tolerant Geobacillus strain. Biotechnology and Bioengineering Volume 102, Issue 5, pp 1377-1386 doi: 10.1002/bit.22181