Directed evolution of AraC for improved compatibility of arabinose- and lactose-inducible promoters
Directed evolution of AraC for improved compatibility of arabinose- and lactose-inducible promoters
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DOI：
10.1128/aem.00791-07
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发表时间：
2007-09-01
影响因子：
4.4
通讯作者：
Keasling, Jay D.
中科院分区：
文献类型：
--
作者：
Lee, Sung Kuk;Chou, Howard H.;Keasling, Jay D.
合成生物系统通常需要多个独立的诱导型启动子以控制几个基因的表达水平;然而，启动子之间的串扰限制了这种能力。在这里，我们证明了AraC的定向进化，以构建阿拉伯糖诱导（P-BAD）系统，该系统与乳糖诱导（P-lac）系统的IPTG（异丙基-β-D-1-硫代半乳糖苷）诱导更相容。构建的系统是10倍更敏感的阿拉伯糖和耐受IPTG显着优于野生型。详细的研究表明，AraC的二聚化结构域和C末端是重要的增加AraC的敏感性，阿拉伯糖。
Synthetic biological systems often require multiple, independently inducible promoters in order to control the expression levels of several genes; however, cross talk between the promoters limits this ability. Here, we demonstrate the directed evolution of AraC to construct an arabinose-inducible (P-BAD) system that is more compatible with IPTG (isopropyl-beta-D-1-thiogalactopyranoside) induction of a lactose-inducible (P-lac) system. The constructed system is 10 times more sensitive to arabinose and tolerates IPTG significantly better than the wild type. Detailed studies indicate that the AraC dimerization domain and C terminus are important for the increased sensitivity of AraC to arabinose.