Improvement of the recombinant phytase expression by intermittent feeding of glucose during the induction phase of methylotrophic yeast Pichia pastoris.

PURPOSE: For growth of methylotrophic yeast, glycerol is usually used as a carbon source. Glucose is used in some cases, but not widely consumed due to strong repressive effect on AOX1 promoter. However, glucose is still considered as a carbon source of choice since it has low production cost and guarantees growth rate comparable to glycerol. RESULTS: In flask cultivation of the recombinant yeast, Pichia pastoris GS115(pPIC9K-appA38M), while methanol induction point(OD600) and methanol concentration significantly affected the phytase expression, glucose addition in induction phase could enhance phytase expression. The optimal flask cultivation conditions illustrated by Response Surface Methodology were 10.37 OD600 induction point, 2.02 h before methanol feeding, 1.16% methanol concentration and 40.36µL glucose feeding amount(for 20 mL culture volume) in which the expressed phytase activity was 613.4 ± 10.2U/mL, the highest activity in flask cultivation. In bioreactor fermentation, the intermittent glucose feeding showed several advantageous results such as 68 h longer activity increment, 149.2% higher cell density and 200.1% higher activity compared to the sole methanol feeding method. These results implied that remaining glucose at induction point might exhibit a positive effect on the phytase expression. CONCLUSION: Glucose intermittent feeding could be exploited for economic phytase production and the other recombinant protein expression by P. pastoris GS115.

Revealing the formation and electrochemical properties of bis(trifluoromethanesulfonyl)imide intercalated graphite with first-principles calculations.

Graphite has been reported to have anion, as well as cation, intercalation capacities as both a cathode and an anode host material for dual ion batteries. In this work, we study the intercalation of bis(trifluoromethanesulfonyl)imide (TFSI) anions from an ionic liquid electrolyte into graphite with first-principles calculations. We build models for TFSI-Cn compounds with systematically increasing graphene sheet unit cell sizes and investigate their stabilities by calculating the formation energy, resulting in the linear decrease of and arrival at the limit of stability. With unit cell sizes identified for stable compound formation, we reveal that the interlayer distance and relative volume expansion ratio of TFSI-Cn increases as we increase the concentration of the TFSI intercalate during the charge process. The electrode voltage is determined to range from 3.8 V to 3.0 V at a specific capacity ranging from 30 mA h g-1 to 54 mA h g-1, in agreement with experiment. Moreover, a very low activation barrier of under 50 meV for TFSI migration, as well as a good electronic conductivity, provide evidence for using these compounds as a promising cathode. Through the analysis of the charge transfer, we clarify the mechanism of TFSI-Cn formation, and reveal new prospects for developing graphite based cathodes.