CommunicationDimerisation of the Glycophorin A Transmembrane Segment in Membranes Probed with the ToxR Transcription Activator

Add time:09/02/2019    Source:sciencedirect.com

Specific interactions between membrane spanning polypeptide segments are important for folding and oligomerisation of integral membrane proteins. Previously the dimerisation of glycophorin A has been shown to depend on interactions between its transmembrane segment by studying chimeric proteins in detergent solution. Here, we examined dimerisation of the glycophorin A transmembrane segment in a natural membrane employing the ToxR transcription activator fromVibrio cholerae. The ToxR protein is integral to the bacterial inner membrane and its activity requires a dimeric state. Therefore, the ToxR protein is suited to monitor quantitative homophilic interactions. We replaced the ToxR transmem brane segment with parts of the glycophorin A transmembrane segment containing the amino acid motif LIxxGVxxGVxxT previously shown to be sufficient for dimerisation in detergent solution. Expression of these chimeric proteins in an indicator strain resulted in strong transcription activation. This is indicative of efficient dimerisation mediated by the glycophorin transmembrane segment inserted into the inner membrane. Analysis of individual point mutants revealed that at least four residues out of this motif are critical for dimer formation in membranes. However, dimerisation of the glycophorin A transmembrane segment appears to be less sensitive to mutations when localised within a natural lipid bilayer compared to measurements in detergent solution. This may be related to a slightly altered structure of the dimer and/or to a higher local concentration and preorientation of the interacting molecules in a membrane. This makes the ToxR system well suited for probing low-affinity interactions between the transmembrane segments of other proteins.

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