Bass S H, Yansura D G
Maxygen, Inc., 515 Galveston Drive, Redwood City, CA 94063, USA.
Mol Biotechnol. 2000 Nov;16(3):253-60. doi: 10.1385/MB:16:3:253.
The Escherichia coli tryptophan (trp) promoter has been used extensively for the high level production of proteins on a small and large scale. This regulated promoter is readily available, relatively easy to turn on, and can be used in essentially any E. coli host background. This article gives a detailed use of the trp promoter including the design of expression vectors, subsequent culture conditions for promoter induction, and, finally, a protocol for the most common way of detecting the newly synthesized protein of interest. Its successful use for heterologous protein expression, however, sometimes requires consideration of parameters other than transcription such as translation initiation, translation elongation, and proteolysis. In this respect we offer guidance in getting through these post-transcriptional problems, which can occur with the use of any promoter.
大肠杆菌色氨酸(trp)启动子已被广泛用于小规模和大规模的蛋白质高水平生产。这种受调控的启动子易于获得,相对容易开启,并且基本上可用于任何大肠杆菌宿主背景。本文详细介绍了trp启动子的使用方法,包括表达载体的设计、启动子诱导的后续培养条件,以及最后检测新合成的目标蛋白质的最常用方法的方案。然而,它在异源蛋白质表达中的成功应用有时需要考虑转录以外的参数,如翻译起始、翻译延伸和蛋白水解。在这方面,我们提供了应对这些转录后问题的指导,这些问题在使用任何启动子时都可能出现。