Thermus Thermophilus as a Thermophilic Model Organism for Biofuels Production

Liu, Jing
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University of Delaware
The potential of using Thermus thermophilus HB8, a thermophilic bacterium, for the production of biofuels was investigated in this work. T. thermophilus is a thermophilic bacterium that thrives naturally at high temperatures, typically around 70-85°C. This bacterium has significant potential in the field of biotechnology because of its unique physical and biochemical properties, especially for the production of low-boiling biofuels. In this project, the growth of Thermus thermophilus HB8 was characterized in batch cultures under defined experimental growth conditions. First, we optimized a defined growth medium to maximize the growth rate of T. thermophilus without the need to use yeast extract in the medium. After that, growth rates were measured under aerobic growth conditions for temperatures ranging between 50°C and 90°C with glucose as the only carbon source. The optimum growth temperature was 80°C and the maximum growth rate of T. thermophilus was 0.28 1/hr, which corresponds to a doubling time of about 2.5 hours. In the growth experiments, T. thermophilus was cultured in custom constructed mini-bioreactors (at 10-mL scale) that were constructed using Hungate tubes and septum caps. For the growth experiments, we used uniformly 13C-labeled [U-13C]glucose as the carbon source and measured 13CO2 production in the off-gas by an on-line mass spectrometer (at m/z 45). With this setup we followed the growth in real-time. In addition to measuring CO2 production, we also measured glucose concentration and optical density at 600 nm (OD600) during the cultures. From all of these measurements, we calculated growth rates for the exponential growth phase. The growth of these cells was then further examined for the potential of producing biofuels by adding various alcohols (ethanol, n-butanol, isobutanol, and propanol) and measuring cell growth. From this data we determined alcohol tolerance for the different potential biofuels. Finally, we applied directed evolution to improve the growth of T. thermophilus HB8. The final adapted cells (termed T. thermophilus JL3 and JL4) displayed higher growth rate, higher tolerance to glucose, higher concentrations of ethanol, and grew to higher cell densities compared to the starter strain T. thermophilus HB8.
Thermus thermophilus , biofuels production