Effect of methanol concentration on the production of human β2-glycoprotein I domain V by a recombinant Pichia pastoris:: A simple system for the control of methanol concentration using a semiconductor Gas Sensor
Effect of methanol concentration on the production of human β2-glycoprotein I domain V by a recombinant Pichia pastoris:: A simple system for the control of methanol concentration using a semiconductor Gas Sensor
复制标题
DOI：
10.1016/s0922-338x(98)80156-6
复制
发表时间：
1998-01-01
期刊：
影响因子：
--
通讯作者：
Suga, KI
中科院分区：
文献类型：
--
作者：
Katakura, Y;Zhang, WH;Suga, KI
甲醇营养型毕赤酵母（Pichia pastoris）由于具有甲醇诱导的AOX 1强启动子，是生产外源蛋白的最佳宿主之一。在外源蛋白的生产阶段，甲醇进料是重要的，因为甲醇不仅诱导蛋白质的生产，而且还为宿主细胞提供能量来源。过量的甲醇抑制宿主细胞的生长，而能量来源不足和/或甲醇饥饿导致生长和生产不良。我们构造了一个简单的甲醇控制系统，由半导体气体传感器和继电器组成。使用该系统，我们研究了甲醇浓度对模型外源蛋白，人β 2-糖蛋白I结构域V的产生的影响。在生产阶段，甲醇浓度保持恒定在1.5、10、17或31 g.l（-1）（+/-5%）。虽然比生长速率和甲醇消耗随着甲醇浓度的增加而降低，但比生产速率增加，表明用于生产的能量与用于细胞生长的能量竞争。因此，我们在生产阶段提供甘油作为额外的能量来源，结果比生产率增加了两倍。该系统操作简单，成本低廉，可用于在毕赤酵母中生产重组蛋白的生物工程研究，目的蛋白的生长和生产依赖于甲醇浓度。
The methylotrophic yeast Pichia pastoris is one of the best hosts for the production of foreign proteins because of the presence of the strong AOX1 promoter induced by methanol. Methanol feeding during the production phase of the foreign proteins is important because methanol not only induces protein production but also provides energy source for the host cells. Excess methanol inhibits the growth of host cells, while an insufficient amount of energy source and/or methanol starvation lead to poor growth and production. We constructed a simple methanol control system consisting of a semiconductor gas sensor and a relay. Using this system, we studied the effect of methanol concentration on the production of a model foreign protein, human beta(2)-glycoprotein I domain V. The methanol concentrations were kept constant at 1.5, 10, 17, or 31 g.l(-1) (+/-5%) during the production phase. Although the specific rates of growth and methanol consumption decreased with increase in the methanol concentration, the specific production rates increased, indicating that the energy for the production competed with that for cell growth. Accordingly, we provided glycerol as an extra energy source during the production phase, with the result that the specific production rate increased two times. Our simple and inexpensive system will help bioengineering studies on the production of recombinant proteins in P. pastoris, the growth and production of objective proteins in which are dependent on the methanol concentration.