文献类型：期刊文献

中文题名：甲醇抑制时重组毕赤酵母发酵的特性研究

英文题名：Fermentation Behaviors of Recombinant Pichia pastoris under Inhibited Methanol Concentration

作者：周祥山[1];范卫民[1];张元兴[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,上海200237

年份：2003

卷号：19

期号：5

起止页码：618

中文期刊名：生物工程学报

外文期刊名：Chinese Journal of Biotechnology

收录：MEDLINE(收录号：15969095);CSTPCD;;Scopus;北大核心:【北大核心2000】；CSCD:【CSCD2011_2012】；PubMed;

语种：中文

中文关键词：甲醇;重组毕赤酵母;发酵;真核表达系统;恒化培养;底物抑制;比生长速率

外文关键词：Pichia pastoris , chemostat culture, substrate inhibition, specific growth rate

摘要：本文对毕赤酵母进行了恒化培养研究。以甲醇为唯一碳源时 ,在稀释率较低时 (D <0 0 4 8h- 1 ) ,连续培养系统操作很稳定。但在稀释率高时 (D >0 0 4 8h- 1 ) ,连续培养系统的定态点不止一个 ,实验不能维持 ,故采用比生长速率恒定的分批流加培养进行研究。结果表明 ,毕赤酵母的生长符合Andrew普遍化底物抑制模型。综合考虑水蛭素的生成、底物的消耗 ,在生产中维持甲醇浓度为限制性浓度 (0 5g L) ,且维持比生长速率为 0 0 2h- 1 时 ,水蛭素Hir6 5的比生成速率达到最大值 0 2mg (g·h)且甲醇的比消耗速率为 0 0 4g (g·h)。
Chemostat culture was performed to characterize the growth, substrate consumption and the hirudin production, and to disclose their interrelations in the fermentation of recombinant Pichia pastoris . The Andrew substrate inhibited growth model is more suitable than Monod model to simulate the growth of Pichia pastoris on methanol. Therefore, two stationary states can be obtained in the continuous culture at a certain dilution rate because of the substrate inhibition on cell growth. The stationary state could be obtained if only the dilution rate not more than 0 048 h -1 in the continuous fermentation. The concentrations of cell, methanol and hirudin were constant after 50 h continuous culture with dilution rate at 0 04 h -1 . However, it could not be obtained when the dilution rate more than 0 048 h -1 because the other stationary point at S >0 048 h -1 is unstable. Therefore, it was found that the cell concentration declined and the methanol concentration increased from 2 9 g/L to 18 1 g/L within 18h at dilution rate 0 06 h -1 . Thus, the fed batch culture with a constant specific growth rate was carried out to disclose the fermentation behavior at high and constant methanol concentration in aid of a methanol sensor. The theoretical maximum specific growth rate, μ m=0 0464 h -1 , was found under critical methanol concentration, S crit =3 1 g/L. The growth of P. pastoris was typically methanol limited at the methanol concentration SS crit . The maximum specific Hir65 production rate q p was obtained at 0 2 mg/(g·h) when methanol concentration and μ were 0 5 g/L and 0 02 h -1 , respectively. The specific Hir65 production rate q p increased with the increase of μ and S at μ <0 02 h -1 , and decreased at μ >0 02 h -1 The specific methanol consumption rate increased with the increase of S when S <5 g/L, but decreased when S >5 g/L. At last, the high Hir65 production rate 0 2 mg/(g·h) was obtained in the fermentation conducted under methanol limited concentration and μ controlled at 0 5 g/L and 0 02 h -1 , respectively, while the specific methanol consumption rate is low only at 0 04 g/(g·h), showing the potential for the strategy of getting high Hir65 production rate at the low consumption of methanol.