Dr. Claire Edwards, a Postdoctoral Research Associate at the BFF, attended the Recent Advancements in Fermentation Technology (RAFT) conference hosted by the Society for Industrial Microbiology and Biotechnology (SIMB). The conference took place October 29 through November 1, 2017 in Bonita Springs, Florida. Claire attended a number of seminars on topics ranging from improvement of oxygen transfer rates to the fermentative production of a heme-bound protein for making meat-less burgers.
She was also able to present some of her work during the first evening’s poster presentation session. Claire has been working on fermentation process optimization for Thermus thermophilus. This genus is known for its ability to grow at high temperatures and you may be familiar with Thermus aquaticus, the strain used to produce Taq polymerase. Enzymes produced by thermophilic organisms such as T. thermophilus are of great interest for basic systems biology experiments, as well as, in the biotechnology industry.
A few of the advantages of working with thermophilic organisms include (1) proteins from extremophiles are typically easier to purify, crystalize and remain more stable over time than mesophilic proteins, (2) thermophilic enzymes have increased resistance to denaturation, and (3) thermophilic enzymes are typically more stable and active in industry relevant conditions. T. thermophilus has the added advantage of being a fast growing strain with high cell yield that constitutively expresses a natural competence apparatus.
The BFF has grown T. thermophilus for the purification of homologous proteins in the past, but had never tried to optimize growth conditions. Claire used the DASGIP parallel bioreactor system to test a number of different conditions to increase OD600 levels during the exponential growth phase. The two factors that had the strongest impact on the growth of T. thermophilus were antifoam addition and glucose concentration.
T. thermophilus growth was inhibited by the addition of all types of antifoam tested. Some antifoams slowed the organism’s growth while others completely stopped it from growing. Decreasing the antifoam addition and adding the antifoam after autoclaving did not affect the culture’s final OD600, but it did increase growth rates and decrease the time the culture spent in lag phase by almost 80%.
Maximum OD600 of T. thermophilus was reached when 6g/L of glucose was added to the growth medium. Lower concentrations (0g/L and 3g/L) and higher concentrations (9g/L) had lower OD600s during the exponential growth phase. There was no benefit to running fed-batch fermentations where the glucose concentration was maintained at approximately 6g/L in the medium.
Comparison of the starting fermentation conditions used historically in the BFF for producing homologous proteins from T. thermophilus with the new conditions showed an increase in OD600 during exponential growth. Fermentations harvested during the exponential growth phase using the optimized conditions produced more cells based on wet weight and more total protein based on a Bradford assay than when the original conditions were used. Furthermore, these optimized conditions decreased harvest time by 45%.