多拷贝毕赤酵母表达猪胰岛素前体的动力学模型

doi:10.11949/j.issn.0438-1157.20151432

化工学报 ›› 2016, Vol. 67 ›› Issue (5): 2015-2021.DOI: 10.11949/j.issn.0438-1157.20151432

多拷贝毕赤酵母表达猪胰岛素前体的动力学模型

陈丽^1,2, 王玥², 郭美锦², 储炬², 庄英萍²

1 江苏省农业科学院国家兽用生物制品工程技术研究中心, 江苏南京 210014;
2 华东理工大学, 上海生物制造技术协同创新中心, 上海 200237

收稿日期:2015-09-10 修回日期:2015-11-19 出版日期:2016-05-05 发布日期:2016-05-05
通讯作者: 郭美锦
基金资助:
农业公益性行业科研专项经费项目(201303046);国家高技术研究发展计划重点项目(2012AA021201);生物反应器工程国重室开放课题资助(2060204)。

Kinetic modelling of porcine insulin precursor (PIP) expressed by multi-copy recombinant Pichia pastoris

CHEN Li^1,2, WANG Yue², GUO Meijin², CHU Ju², ZHUANG Yingping²

1 National Research Center of Engineering and Technology for Veterinary Biologicals, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, Jiangsu, China;
2 Shanghai Collaborative Innovation Center for Biomanufacturing Technology, East China University of Science and Technology, Shanghai 200237, China

Received:2015-09-10 Revised:2015-11-19 Online:2016-05-05 Published:2016-05-05
Supported by:
supported by the Special Fund for Agro-scientific Research in the Public Interest(201303046), the National High Technology Research and Development Program of China (2012AA021201), and the Open Project Program of State Key Laboratory of Bioreactor Engineering (2060204).

摘要/Abstract

摘要：

以携带多拷贝猪胰岛素前体(PIP)基因的毕赤酵母(Pichia pastoris)为供试菌株,对分批补料发酵PIP合成动力学模型进行研究。建立了不同拷贝数毕赤酵母发酵合成PIP的菌体生长、产物合成和底物消耗的动力学模型,并通过Origin8.0软件对模型参数进行了非线性拟合。根据动力学模型结果发现,随着拷贝数的增加与菌体生长相关的产物系数和细胞生长代谢系数k₁绝对值不断增加,12拷贝时PIP表达量达到最高,说明只有进一步促进高拷贝菌株细胞生长并减少其代谢负担才能更有效地提高产物的生成速率。同时表明:预测值与实验值有良好的拟合性,所建模型能较好地反映分批补料发酵过程PIP的合成。

关键词: 重组毕赤酵母, 猪胰岛素前体表达, 拷贝数, 动力学模型

Abstract:

Kinetic modelling of recombinant Pichia pastoris harboring multiple porcine insulin precursor (PIP) gene dosages was studied when the cells were grown in the fed-batch culture. The key parameters of this kinetics were estimated, including specific cell growth rate (h^-1), PIP production rate (g·g^-1·h^-1) and substrate consumption rate (g·g^-1·h^-1) with nonlinear curve fit by Origin8.0. The results showed that both growth-associated production coefficient ( ) and growth-associated metabolism coefficient (k₁) increased with increasing copy numbers. The expression level of PIP reached the highest at the copy number of 12. These results suggested that rapid growth and lower metabolic burden of a high copy number effectively improved the production rate of target proteins. Furthermore, the predicted values based on the established kinetic model were in good agreement with the experimental data, indicating that the kinetic model could be used to describe recombinant PIP production process in fed-batch fermentation mode.

Key words: recombinant Pichia pastoris, porcine insulin precursor expression, gene dosage, kinetic model

中图分类号:

TQ464.7

陈丽, 王玥, 郭美锦, 储炬, 庄英萍. 多拷贝毕赤酵母表达猪胰岛素前体的动力学模型[J]. 化工学报, 2016, 67(5): 2015-2021.

CHEN Li, WANG Yue, GUO Meijin, CHU Ju, ZHUANG Yingping. Kinetic modelling of porcine insulin precursor (PIP) expressed by multi-copy recombinant Pichia pastoris[J]. CIESC Journal, 2016, 67(5): 2015-2021.

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多拷贝毕赤酵母表达猪胰岛素前体的动力学模型

Kinetic modelling of porcine insulin precursor (PIP) expressed by multi-copy recombinant Pichia pastoris

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