Plantz Bradley A, Sinha Jayanta, Villarete Lorelie, Nickerson Kenneth W, Schlegel Vicki L
School of Biological Sciences, Beadle Center, University of Nebraska, Lincoln, NE 68583-0666, USA.
Appl Microbiol Biotechnol. 2006 Sep;72(2):297-305. doi: 10.1007/s00253-005-0271-8. Epub 2006 Feb 22.
A growth-associated model was applied to the production of recombinant ovine interferon-tau (rOvIFN-tau) with Pichia pastoris for the purpose of manufacturing preclinical and clinical active material. This model predicts that product yields will be the greatest when the specific growth of the culture is maintained at a steady and optimal rate. However, rOvIFN-tau yields did not meet the expected linear model but most closely corresponded to a polynomial relationship. After transitioning from glycerol to methanol, product accumulated for 31-45 h, and then the yield decreased. This production shift, which has been termed decoupling, was clearly related to time on methanol and not culture density. It was determined that a correlation exists between the decoupling point and a drop in energy state of the cell when expressing beta-galactosidase. By assigning decoupling as a constraint that limits productivity and by reformulating the growth medium, the time prior to decoupling increased to 46.8+/-2.4 h, product yield improved for rOvIFN-tau from 203 to 337 mg l(-1), and the coefficient of variation for yield decreased from 67.9 to 23.3%. A robust and stable fermentation process was realized, resulting in a 210% improvement in total yield from 557+/-357 to 1,172+/-388 mg.
为了生产临床前和临床活性材料，将生长相关模型应用于毕赤酵母生产重组羊干扰素 - τ（rOvIFN - τ）。该模型预测，当培养物的比生长速率保持在稳定且最佳的水平时，产品产量将达到最大。然而，rOvIFN - τ的产量并未符合预期的线性模型，而是最接近多项式关系。从甘油过渡到甲醇后，产物积累31 - 45小时，然后产量下降。这种生产转变被称为解耦，显然与甲醇培养时间有关，而不是与培养密度有关。已确定在表达β - 半乳糖苷酶时，解耦点与细胞能量状态的下降之间存在相关性。通过将解耦设定为限制生产力的约束条件，并重新配制生长培养基，解耦前的时间增加到46.8±2.4小时，rOvIFN - τ的产品产量从203毫克/升提高到337毫克/升，产量变异系数从67.9%降至23.3%。实现了稳健且稳定的发酵过程，总产量从557±357毫克提高到1172±388毫克，提高了210%。
