Effects of glycine substitutions on the structure and function of gramicidin a channels
by Jordan, J. B.; Shobana, S.; Andersen, O. S.; Hinton, J. F.
Tryptophan residues often are found at the lipid-aqueous interface region of membrane-spanning proteins, including ion channels, where they are thought to be important determinants of protein structure and function. To better understand how Trp residues modulate the function of membrane-spanning channels, we have examined the effects of Trp replacements on the structure and function of gramicidin A channels. Analogues of gramicidin A in which the Trp residues at positions 9, 11, 13, and 15 were sequentially replaced with Gly were synthesized, and the three-dimensional structure of each analogue was determined using a combination of two-dimensional NMR techniques and distance geometry-simulated annealing structure calculations. Though Trp -> Gly substitutions destabilize the beta(6.3)-helical gA channel structure, it is possible to determine the structure of analogues with Trp -> Gly substitutions at positions 11, 13, and 15, but not for the analogue with the Trp -> Gly substitution at position 9. The Gly(11)-, Gly(13)-, and Gly(15)-gA analogues form channels that adopt a backbone fold identical to that of native gramicidin A, with only small changes in the side chain conformations of the unsubstituted residues. Single-channel current measurements show that the channel function and lifetime of the analogues are significantly affected by the Trp -> Gly replacements. The conductance variations appear to be caused by sequential removal of the Trp dipoles, which alter the ion-dipole interactions that modulate ion movement. The lifetime variations did not appear to follow a clear pattern.
- Journal
- Biochemistry
- Volume
- 45
- Issue
- 47
- Year
- 2006
- Start Page
- 14012-14020
- URL
- https://dx.doi.org/10.1021/bi061560z
- ISBN/ISSN
- 1520-4995; 0006-2960
- DOI
- 10.1021/bi061560z