NMR Study of Gramicidin Cation Transport Across and Integration into a Lipid Membrane

Abstract

Gramicidin is a 15 amino acid linear polypeptide which forms cation conducting channels in lipid membranes (Meyers and Haydon, 1972; Anderson, 1983). The gramicidin channel in a lipid bilayer membrane is shown in Figure 1. The amino acid sequence of gramicidin is: formyl-L-Val1-Gly2-L-Ala3-D-Leu4-L-Ala5-D-Val6- L-Val7-D-Val8-L-Trp9-D-Leu10-L-Trp11-D-Leu12-L-Trp13-D-Leu14-L-Trp15-ethanolamine The channel consists of two monomeric beta helices joined by hydrogen bonds at their NH2 terminal ends. The dimmer’s length is 26 A and forms a 4 A pore. The three naturally occurring analogs Gramicidin A, B, and C differ by one amino acid, tryptophan, phenylalanine, and tyrosine respectively, at position 11 (Sarge amp; Witkop, 1965). Here we report the preliminary results of the Na-23 NMR investigation of the thermal integration of gramicidin analogs into large unilamellar vesicles under ionic equilibrium conditions. This technique is potentially useful for exploring the effect of single amino acid substitution in gramicidin on the rate of incorporation in and formation of a viable channel in a lipid membrane. The effect of different lipids on the incorporation process can also be studied using this technique. The only requirement for a peptide to be studied in this way is for the peptide to conduct ions at a rate sufficient to cause a change in the linewidth of the NMR signal of the conducting ion. The incorporation of gramicidin into lysolecithin micelles has been investigated previously by C-13 NMR (Spisni et al., 1979). This technique, however, is less amiable to studying the rate of incorporation. The incorporation of gramicidin into lysophosphatidylcholine micelles was also studied using fluorescence measurements (Cavatorta et al., 1982). However, with our system the peptide does not have to have a fluorescence or absorbance spectrum.