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中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志
中国生物工程杂志  2015, Vol. 35 Issue (11): 23-28    DOI: 10.13523/j.cb.20151104
研究报告     
ste3与ste4基因双敲除对依博素生物合成的影响
白利平, 姜蓉, 郭连宏, 张洋, 李元
中国医学科学院/北京协和医学院医药生物技术研究所 北京 100050
The Effects of ste3 and ste4 Genes Double Disruption in Ebosin Biosynthesis
BAI Li-ping, JIANG Rong, GUO Lian-hong, ZHANG Yang, LI Yuan
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100050, China
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摘要：

目的:以往研究已确定链霉菌胞外多糖依博素的生物合成基因簇(ste), 生物信息学分析基因簇中 ste3 和ste4 编码糖转运相关膜蛋白,现研究分析 ste3 和ste4 与依博素生物合成的相关性。方法:通过基因同源重组双交换获得 ste3 和ste4 双基因缺失突变株,经Southern 杂交验证后,对该菌株进行了基因互补研究。分离提取各菌株发酵液的胞外多糖,并计算产量。结果:双基因缺失株产生的胞外多糖依博素与野生株相比,产量从319mg/L降低至153mg/L,平均产量降低52%以上;基因互补后,依博素平均产量上升到299mg/L,基本恢复至野生株依博素产量水平。结论:本研究首次阐明了编码糖转运相关膜蛋白基因 ste3 和ste4 在依博素的生物合成中起重要作用,为研究链霉菌139的初级代谢和次级代谢之间的相关性奠定了基础。
关键词： 链霉菌;  ste3 ste4;  基因双敲除;  依博素    
Abstract:

Objective: The biosynthesis cluster (ste) of a novel exololysaccharide called Ebosin producing by Streptomyces had been identified previously,that ste3 and ste4 maybe encode a membrane spanning protein probably for sugar import on the basis of bioinformatics analysis. This work studied If ste3 and ste4 were involved in Ebosin biosynthesis. Methods: the ste3 and ste4 genes were disrupted together with a double crossover via homologous recombination. The mutant strain was identified by southern blot and gene complementation also performed. The production of Ebosin through isolating it from wild type, mutant and complementation was analyzed. Results: Ebosin isolated from the supernatants of fermentation cultures of the mutant strain was 153mg/L, remarkably lower than that of the wild-type strain (319mg/L). Complementation of the mutant strain resulted in a recovery of the EPS production (299mg/L) comparing with the wild-type strain. Conclusion: This research firstly elucidate that genes ste3 and ste4 encoding a membrane spanning protein involved in Ebosin biosynthesis and lay the foundation for studying the relationship between primary and second metabolism of Streptomyces sp. 139.
Key words: ste3 ste4    Ebosin    Streptomyces    Gene double disruption
收稿日期: 2015-06-25 出版日期: 2015-11-25
ZTFLH:  Q78  
基金资助:

国家自然科学基金(31070086),北京市自然科学基金(5092020),北京协和医学院协和青年科研基金(3332014009,3332015166),中央级公益性科研院所基本科研业务专项(IMBF2015011)资助项目
通讯作者: 白利平     E-mail: lipingbai1973@163.com
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引用本文:

白利平, 姜蓉, 郭连宏, 张洋, 李元. ste3与ste4基因双敲除对依博素生物合成的影响[J]. 中国生物工程杂志, 2015, 35(11): 23-28.

BAI Li-ping, JIANG Rong, GUO Lian-hong, ZHANG Yang, LI Yuan. The Effects of ste3 and ste4 Genes Double Disruption in Ebosin Biosynthesis. China Biotechnology, 2015, 35(11): 23-28.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20151104    或    https://manu60.magtech.com.cn/biotech/CN/Y2015/V35/I11/23

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