Artificial Caprolactam-Specific Riboswitch as an Intracellular Metabolite Sensor
Artificial Caprolactam-Specific Riboswitch as an Intracellular Metabolite Sensor
复制标题
DOI：
10.1021/acssynbio.8b00452
复制
发表时间：
2019-06-01
影响因子：
4.7
通讯作者：
Jung, Gyoo Yeol
中科院分区：
文献类型：
--
作者：
Jang, Sungyeon;Jang, Sungho;Jung, Gyoo Yeol
己内酰胺是合成尼龙-6的单体，最近开发了一种用于生物基生产尼龙-6的重组微生物菌株。己内酰胺的细胞内生物传感器可以促进重组微生物菌株的高通量代谢工程。由于重组菌株中己内酰胺和戊内酰胺的混合生产，己内酰胺生物传感器应该对己内酰胺具有高度特异性。然而，尚未报道高度特异性的己内酰胺传感器。在这里，我们开发了一种人工核糖开关，专门响应己内酰胺。该核糖开关使用偶联的体外体内选择策略与从体外选择获得的RNA适体的异质池来制备，以构建用于体内选择的核糖开关文库。己内酰胺核糖开关成功区分己内酰胺和戊内酰胺。此外，在50 mM己内酰胺的存在下，核糖开关被激活3.36倍。这种核糖开关使细胞生长的己内酰胺依赖性控制成为可能，这将有助于提高己内酰胺的生产，是代谢工程的一个有价值的工具。
Caprolactam is a monomer used for the synthesis of nylon-6, and a recombinant microbial strain for biobased production of nylon-6 was recently developed. An intracellular biosensor for caprolactam can facilitate high throughput metabolic engineering of recombinant microbial strains. Because of the mixed production of caprolactam and valerolactam in the recombinant strain, a caprolactam biosensor should be highly specific for caprolactam. However, a highly specific caprolactam sensor has not been reported. Here, we developed an artificial riboswitch that specifically responds to caprolactam. This riboswitch was prepared using a coupled in vitro in vivo selection strategy with a heterogeneous pool of RNA aptamers obtained from in vitro selection to construct a riboswitch library used in in vivo selection. The caprolactam riboswitch successfully discriminated caprolactam from valerolactam. Moreover, the riboswitch was activated by 3.36-fold in the presence of 50 mM caprolactam. This riboswitch enabled caprolactam-dependent control of cell growth, which will be useful for improving caprolactam production and is a valuable tool for metabolic engineering.