An extended ToxR POSSYCCAT system for positive and negative selection of self-interacting transmembrane domains

Add time:08/31/2019    Source:sciencedirect.com

Assay systems based on the ToxR protein are widely used to investigate interaction of transmembrane domains that come from natural proteins or are isolated from combinatorial libraries. The principle of this method is that self-interaction of any given transmembrane domain, which is expressed within a ToxR chimeric protein, drives ToxR–ToxR assembly in a bacterial inner membrane. In current versions of the system, ToxR–ToxR interaction drives transcription activation of the cholera toxin (ctx) promoter and thereby induces expression of downstream reporter genes in appropriately constructed bacterial strains. Here, we describe the application of other known ToxR-regulated promoters. We show that interacting transmembrane domains also promote ToxR-driven activation of the ompU promoter. Conversely, these interactions efficiently repress transcription from the constitutively active ompT promoter. We present novel Escherichia coli strains whose chromosomes harbor fusions of ompU or ompT promoters with different reporter genes. Depending on the used promoter, self-interaction of transmembrane domains induces or represses reporter enzyme expression in these cells. These strains extend current applications of the ToxR protein and may find use in mapping transmembrane helix–helix interfaces and selection of transmembrane domains with medium affinities.

We also recommend Trading Suppliers and Manufacturers of ToxR protein (cas 114265-38-2). Pls Click Website Link as below: cas 114265-38-2 suppliers

Prev:Multi-Tox: Application of the ToxR-transcriptional reporter assay to the study of multi-pass protein transmembrane domain oligomerization
Next:Structural studies on Vibrio cholerae ToxR periplasmic and cytoplasmic domains)