Reimer Artur, Yagur-Kroll Sharon, Belkin Shimshon, Roy Shantanu, van der Meer Jan Roelof
Department of Fundamental Microbiology, University of Lausanne, Bâtiment Biophore, Quartier UNIL-Sorge 1015 Lausanne, Switzerland.
Department of Plant and Environmental Sciences, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
Sci Rep. 2014 Jul 9;4:5626. doi: 10.1038/srep05626.
Bioreporter bacteria, i.e., strains engineered to respond to chemical exposure by production of reporter proteins, have attracted wide interest because of their potential to offer cheap and simple alternative analytics for specified compounds or conditions. Bioreporter construction has mostly exploited the natural variation of sensory proteins, but it has been proposed that computational design of new substrate binding properties could lead to completely novel detection specificities at very low affinities. Here we reconstruct a bioreporter system based on the native Escherichia coli ribose binding protein RbsB and one of its computationally designed variants, reported to be capable of binding 2,4,6-trinitrotoluene (TNT). Our results show in vivo reporter induction at 50 nM ribose, and a 125 nM affinity constant for in vitro ribose binding to RbsB. In contrast, the purified published TNT-binding variant did not bind TNT nor did TNT cause induction of the E. coli reporter system.
生物报告菌，即通过产生报告蛋白来响应化学物质暴露而设计的菌株，因其有潜力为特定化合物或条件提供廉价且简单的替代分析方法而引起了广泛关注。生物报告菌的构建大多利用了传感蛋白的自然变异，但有人提出，通过计算设计新的底物结合特性，有可能在非常低的亲和力下实现全新的检测特异性。在此，我们基于天然的大肠杆菌核糖结合蛋白RbsB及其一种经计算设计的变体构建了一个生物报告系统，据报道该变体能够结合2,4,6-三硝基甲苯（TNT）。我们的结果显示，在50 nM核糖浓度下，报告基因在体内被诱导，体外核糖与RbsB结合的亲和常数为125 nM。相比之下，纯化后的已发表的TNT结合变体既不结合TNT，TNT也未引起大肠杆菌报告系统的诱导。
