恒细胞密度发酵策略提高重组毕赤酵母生产碱性果胶酶的表达效率

Enhancing expression efficiency of alkaline polygalacturonate lyase by constant cell concentration culture of the recombinant Pichia pastoris

摘要:

为了提高重组毕赤酵母生产碱性果胶酶 (Alkaline polygalacturonate lyase，PGL) 的比速率，开发了一种新的恒细胞密度发酵策略。通过不同的甲醇流加方式，实现发酵过程细胞密度的合理控制。实验结果表明：控制细胞密度为75 g/L的策略为最优，最终单位发酵液体积生产强度和单位菌体生产强度为6.11 U/(mL·h) 和81.5 U/(g·h)，分别比传统高密度发酵提高了42.1％和191.2％，最终PGL酶活为441.9 U/mL。此外，该策略还具有提高细胞活性和降低蛋白酶降解作用

Abstract:

In order to enhance the alkaline polygalacturonate lyase (PGL) productivity by Pichia pastoris, we developed a constant cell concentration culture strategy by methanol feeding (called as CCCM culture) used in the continuous cultures. We controlled reasonable cell concentrations in the bioprocess by different strategies of methanol feeding. Using this CCCM culture with DCW 75 g/L, we significantly enhanced the PGL productivity (QV) and the average specific enzyme production rate (QX) of PGL to 6.11 U/(mL·h) and 81.5 U/(g·h), increased by 42.1% and 191.2% than the fed-batch culture with high cell density, respectively. The final PGL activity was 441.9 U/mL. Moreover, the extracellular protease concentration is 1.9 mg/L and the cell viability is more than 94% after 120 hour induction. The results show that this new strategy is advantageous in reducing proteolytic degradation and enhancing cell viability.