重组大肠杆菌生产类人胶原蛋白Ⅱ的发酵pH优化-陕西师范大学学报期刊社网站

陕西师范大学学报（自然科学版）

生命科学

重组大肠杆菌生产类人胶原蛋白Ⅱ的发酵pH优化

王维卓1,2，范代娣1,2,3*，骆艳娥1,2,3，马晓轩1,2,3，朱晨辉1,2,3，郭佳庆1,2

（1 西北大学化工学院； 2 陕西省生物材料与发酵工程技术研究中心；3 陕西省可降解生物医用材料重点实验室, 陕西西安 710069）

王维卓，男，硕士研究生,主要研究方向为发酵过程优化.* 通信作者：范代娣,女,教授,博士研究生导师.E-mail:fandaidi@nwu.edu.cn.

摘要:

为了实现类人胶原蛋白(HLCⅡ)的规模化生产,对重组大肠杆菌BL21 37在30 L发酵罐中发酵生产类人胶原蛋白Ⅱ过程中的关键参数pH进行了系统的研究.将发酵过程中pH控制在60~78之间,监测pH对细胞生长以及类人胶原蛋白Ⅱ的产量的影响,从而确定出合理的pH控制策略增加类人胶原蛋白Ⅱ的产量.由实验数据可得,在pH75条件下细胞浓度达到最大,为685 g／L,但HCLⅡ的浓度为85 g／L.在pH68条件下,HLCⅡ浓度达到最大,为105 g／L,但其细胞浓度只有609 g／L.同时,对发酵过程中代谢副产物有机酸的含量也进行了监测,随着发酵过程pH值的升高,总有机酸的量也相应的增加,当发酵过程pH控制在78时,总有机酸达到最大,为1561 g／L.为了提高目标产物的产量,采用pH两阶段控制策略控制发酵,最终类人胶原蛋白Ⅱ浓度可达到113 g／L,较恒pH条件下提高了761％.

关键词：

类人胶原蛋白Ⅱ；重组大肠杆菌； pH两阶段控制；有机酸

收稿日期：

2009-12-11

中图分类号：

Q815

文献标识码：

文章编号：

1672-4291(2010)05-0072-05

基金项目：

国家“863”计划基金资助项目（2006AA02Z246;2007AA03Z456）；国家自然科学基金资助项目（20776119）；陕西省教育厅科学研究发展计划基金资助项目(07JK417)

Doi:

Optimal Strategy of pH Control in the Production ofHLCⅡ by Recombinant Escherichia coli

WANG Wei-zhuo1，2, FAN Dai-di1，2，3*, LUO Yan-e1， 2,3, MA Xiao-xuan1，2,3, ZHU Chen-hui1，2,3, GUO Jia-qing1，2

（1 College of Chemical Engineering, Northwest University；2 Shaanxi R＆D Center of Biomaterials and Fermantation Engeering；3 Shaanxi Key Laboratory of Degradable Biomedical Materials, Xi′an 710069,Shaanxi,China）

Abstract:

To achieve large-scale production of HLC Ⅱ, the key parameter pH in production process of human-like collagen by recombinant Escherichia coli BL21 37 was systematicly investigated in a 30 L fermentation tank with triplex turbine impeller. The effect of pH ranging from 60 to 78 on cell content and HLC Ⅱ production during fermentation process was studied, and an optimal pH strategy was determined for increase of HLC Ⅱ yield. From experimental data, the cell growth and collagen synthesis were significantly affected by pH. An optimal pH of 75 for cell growth was obtained with cell density at 685 g／L, the concentration of HLC Ⅱ was 85 g／L; while an optimal pH of 68 for HLC Ⅱ accumulation was obtained with concentration of HLC Ⅱ at 105 g／L, and cell density was 609 g／L. Additionally, the total organic acid of fermentation process was monitored. The concentration of organic acid increased with pH, and the highest concentration of organic acid, 1561 g／L, was observed when pH was kept constant at 78 during the process of fermentation. Based on the results, a pH shift method was developed to gain higher concentration of HCL Ⅱ which reached to 113 g／L, increased by 761％ as compared with constant pH fermentation.

KeyWords:

Human-like collagen Ⅱ; recombinant escherichia coli; pH shift control; organic acid