High dissolved oxygen tension enhances heterologous protein expression by recombinant Pichia pastoris
High dissolved oxygen tension enhances heterologous protein expression by recombinant Pichia pastoris
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DOI：
10.1016/s0032-9592(02)00280-7
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发表时间：
2003-03-28
影响因子：
4.4
通讯作者：
Lee, EK
中科院分区：
文献类型：
--
作者：
Lee, CY;Lee, SJ;Lee, EK
在DO-stat补料分批发酵中，采用不同设定值和带宽的DO控制带，研究了DO张力对毕赤酵母系统表达外源蛋白的影响。模型蛋白是弹性蛋白酶抑制肽（EIP），它对人中性粒细胞弹性蛋白酶具有很强的抑制活性，具有很高的特异性。在醇加氧酶I去阻遏阶段，通过保持DO控制带较窄而更频繁地补加甘油导致更高的细胞生长速率（0.10 h（-1））。在甲醇诱导阶段，保持较高的DO设定点并允许更频繁的补料显著增强了EIP表达。通过将较低的设定点从10%空气饱和度提高到30%空气饱和度，EIP表达滴度平均增加约3倍至846 mg l（-1）。比表达滴度和甲醇消耗速率均提高了约3倍，表明较高的DO张力刺激了甲醇利用途径，提高了表达效率。为了以更高的DO设定点操作DO-stats，可以从常规空气产生高达38%的氧分压的富氧膜系统被用作由空气压缩机和纯氧瓶组成的传统气体混合系统的替代氧气供应器。(C)2002爱思唯尔科技有限公司版权所有。
The effect of dissolved oxygen (DO) tension on heterologous protein expression by the Pichia system was investigated by applying DO control bands of different set points and bandwidths in the DO-stat fed-batch fermentation. The model protein was elastase inhibiting peptide (EIP) that showed strong inhibitory activity against human neutrophil elastase with very high specificity. In the alcohol oxygenase I derepression phase, more frequent glycerol feeding by keeping the DO control band narrower resulted in a higher cell growth rate (0.10 h(-1)). In the methanol induction phase, maintaining a higher DO set point and allowing more frequent feeding significantly enhanced EIP expression. The EIP expression titre was increased about threefold to 846 mg l(-1) on average by raising the lower set point from 10 to 30% air-saturation. Both specific expression titre and methanol consumption rates were increased about threefold, which indicated that higher DO tension stimulated the methanol utilization pathway and improved the expression efficiency. To operate the DO-stats with higher DO set points, an oxygen enriching membrane system that could generate up to 38% oxygen partial pressure from the regular air was used as an alternative oxygen supplier to the traditional gas-mixing system consisting of an air compressor and a pure oxygen cylinder. (C) 2002 Elsevier Science Ltd. All rights reserved.