Yu P, San K Y
Chemical Engineering Department, Rice University, Houston, Texas 77251-1892.
Biotechnol Prog. 1993 Nov-Dec;9(6):587-93. doi: 10.1021/bp00024a004.
Continuous production of extracellular recombinant protein using a previously reported periplasmic protein-release system was successfully demonstrated. The performance of the protein-releasing system was studied using two types of reactor configuration: a conventional chemostat and a chemostat with a cell-recycle system. The constitutively expressed alpha-amylase was used as a model protein. In the chemostat experiments, the total alpha-amylase activity increases with decreasing dilution rate--from 135.5 units/mL at a dilution rate of 0.05 h-1 to 2.06 units/mL at a dilution rate of 0.491 h-1. However, the percentage release of alpha-amylase remained relatively constant, at approximately 29%, in the range of dilution rates investigated. The reactor productivity of the target protein reached a maximum of 8.14 units/mL/h at a dilution rate of 0.1 h-1. Application of the releasing system for the continuous production of cell-free recombinant proteins in a cell-recycle fermentor was also studied. The reactor performance is significantly better than that of either the chemostat or the batch mode. A total productivity of alpha-amylase at 13.2 units/mL/h with 42% recovery of proteins in the cell-free form was obtained at a dilution rate of 0.397 h-1 and a bleeding rate of 9.8%.
利用先前报道的周质蛋白释放系统成功实现了细胞外重组蛋白的连续生产。使用两种类型的反应器配置研究了蛋白释放系统的性能：传统的恒化器和带有细胞循环系统的恒化器。组成型表达的α-淀粉酶用作模型蛋白。在恒化器实验中，总α-淀粉酶活性随稀释率降低而增加——从稀释率为0.05 h-1时的135.5单位/毫升降至稀释率为0.491 h-1时的2.06单位/毫升。然而，在所研究的稀释率范围内，α-淀粉酶的释放百分比保持相对恒定，约为29%。目标蛋白的反应器生产力在稀释率为0.1 h-1时达到最大值8.14单位/毫升/小时。还研究了该释放系统在细胞循环发酵罐中连续生产无细胞重组蛋白的应用。该反应器的性能明显优于恒化器或分批模式。在稀释率为0.397 h-1和排料率为9.8%时，α-淀粉酶的总生产力为13.2单位/毫升/小时，无细胞形式的蛋白回收率为42%。
