文献类型：期刊文献

英文题名：Reverse biological engineering of hrdB to enhance the production of avermectins in an industrial strain of Streptomyces avermitilis

作者：Zhuo, Ying[1,4];Zhang, Wenquan[1];Chen, Difei[1];Gao, Hong[1];Tao, Jun[1,4];Liu, Mei[1];Gou, Zhongxuan[1];Zhou, Xianlong[1];Ye, Bang-Ce[2];Zhang, Qing[3];Zhang, Siliang[2];Zhang, Li-Xin[1]

机构：[1]Chinese Acad Sci, Inst Microbiol, Key Lab Pathogen Microbiol & Immunol, Beijing 100190, Peoples R China;[2]E China Univ Sci & Technol, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China;[3]Hebei Veyong Biochem Co Ltd, Shijiazhuang 050031, Hebei, Peoples R China;[4]Chinese Acad Sci, Grad Univ, Beijing 100190, Peoples R China

年份：2010

卷号：107

期号：25

起止页码：11250

外文期刊名：PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000279058000020)】；

基金：The authors are very grateful for the technical support provided by Huimin Yu from Tsinghua University. They also thank Elizabeth Ashforth, Huarong Tan, Zixin Deng, Linquan Bai, Zhiheng Liu, Jibin Sun, Keqian Yang, Xiaopeng Jia, and Yaqiao Li for critical reading and helpful discussions. This work was supported in part by grants from National 863 Project (2006AA09Z402, 2007AA09Z443), Key Project for International Cooperation (2007DFB31620), National Natural Science Foundation of China (30560001, 30600001, 30700015), National Key Technology R&D Program (2007BAI26B02), the CAS Pillar Program (KSCX2-YW-R-164), and Important National Science & Technology Specific Projects (2008ZX09401-05, 2009ZX09302-004). L.-X.Z. received funding from the Hundred Talents Program.

语种：英文

外文关键词：precision engineering; RNA polymerase; overproduction

摘要：Avermectin and its analogues are produced by the actinomycete Streptomyces avermitilis and are widely used in the field of animal health, agriculture, and human health. Here we have adopted a practical approach to successfully improve avermectin production in an industrial overproducer. Transcriptional levels of the wildtype strain and industrial overproducer in production cultures were monitored using microarray analysis. The avermectin biosynthetic genes, especially the pathway-specific regulatory gene, aveR, were up-regulated in the high-producing strain. The upstream promoter region of aveR was predicted and proved to be directly recognized by sigma(hrdB) in vitro. A mutant library of hrdB gene was constructed by error-prone PCR and selected by high-throughput screening. As a result of evolved hrdB expressed in the modified avermectin high-producing strain, 6.38 g/L of avermectin B1a was produced with over 50% yield improvement, in which the transcription level of aveR was significantly increased. The relevant residues were identified to center in the conserved regions. Engineering of the hrdB gene can not only elicit the overexpression of aveR but also allows for simultaneous transcription of many other genes. The results indicate that manipulating the key genes revealed by reverse engineering can effectively improve the yield of the target metabolites, providing a route to optimize production in these complex regulatory systems.