Identification of a translocated protein segment in a voltage-dependent channel
- 1 September 1994
- journal article
- Published by Springer Nature in Nature
- Vol. 371 (6493), 158-161
- https://doi.org/10.1038/371158a0
Abstract
Voltage-gated channels undergo a conformational change in response to changes in transmembrane voltage. Here we use site-directed biotinylation to create conformation-sensitive sites on colicin Ia, a bacteriocidal protein that forms a voltage-sensitive membrane channel, which can be monitored by electrophysiological methods. We investigated a model of gating developed for the partly homologous colicin E1 that is based on the insertion of regions of the protein into the membrane in response to cis-positive voltages. Site-directed cysteine mutagenesis, followed by chemical modification, was used to attach a biotin molecule covalently to a series of unique sites on colicin Ia. The modified protein was incorporated into planar lipid membranes, where the introduced biotin moiety served as a site to bind the water-soluble protein streptavidin, added to one side of the membrane or the other. Our results show that colicin gating is associated with the translocation across the membrane of a segment of the protein of at least 31 amino acids.Keywords
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