期刊
ENZYME AND MICROBIAL TECHNOLOGY
卷 68, 期 -, 页码 33-42出版社
ELSEVIER SCIENCE INC
DOI: 10.1016/j.enzmictec.2014.10.006
关键词
Pichia pastoris; Heterologous protein secretion; Glucose oxidase; Modular engineering
资金
- National High Technology Research and Development Program of China (863 Program) [2011AA100905]
- Major State Basic Research Development Program of China (973 Program) [2013CB733902]
- National Natural Science Foundation of China [31470160]
- China Postdoctoral Science Foundation [114957, 2013M540538]
- Synergetic Innovation Center of Food Safety and Nutrition
- Natural Science Foundation of Jiangsu Province [BK2012553]
Limitations in protein production and secretion have been attributed to the inefficient folding rate of overexpressed proteins and the cellular response to the presence of overexpressed proteins in the endoplasmic reticulum (ER). In this study, we improved the yield of glucose oxidase (GOD) by manipulating genes involved in protein folding machinery and abnormal folding stress responses. First, genes with folding and secretion functions were used to modulate the folding rate of GOD in the ER and its secretion level in the cytoplasm. Next, the potential benefits of the ERAD elements were determined. Cellular resistance to ER derived stress was then strengthened by overexpressing the stress response gene GCN4. Furthermore, a module combination strategy, which co-expressed the SEC53, CNE1 and GCN4 genes, was employed to construct the Pichia pastoris strain S17. This increased the yield of GOD to 21.81 g/L, with an activity of 1972.9 U/mL, which were 2.53- and 5.11-fold higher, respectively, than the control strain. The work described here improved GOD production significantly, and the strategies employed in this study provide novel information for the large-scale production of heterologous proteins. (C) 2014 Elsevier Inc. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据