Gramicidin tryptophans mediate formamidinium-induced channel stabilization.
Compared with alkali metal cations, formamidinium ions stabilize the gramicidin A channel molecule in monoolein bilayers (Seoh and Busath, 1993a). A similar effect is observed with N-acetyl gramicidin channel molecules in spite of the modified forces at the dimeric junction (Seoh and Busath, 1993b). Here we use electrophysiological measurements with tryptophan-to-phenylalanine-substituted gramicidin analogs to show that the formamidinium-induced channel molecule stabilization is eliminated when the four gramicidin tryptophans are replaced with phenylalanines in gramicidin M-. This suggests that the stabilization is mediated by the tryptophan side chains. Tryptophan residues 9, 13, and 15 must cooperate to produce the effect because replacement of any one of the three with phenylalanine significantly reduces stabilization; replacement of Trp-11 with phenylalanine causes negligible decrease in stabilization. In addition, formamidinium-related current-voltage supralinearity and open-channel noise are absent with gramicidin M-. When the lipid bilayer was formed with monoolein ether rather than monoolein ester, the channel lifetimes were reduced markedly and, at low voltage and relative to those in KCl solution, were decreased by a factor of 2, whereas the open-channel noise was unaffected and the current-voltage relation was only modestly affected. These results suggest that formamidinium modifies the state of the tryptophan side chains, which, in turn, affects channel lifetime, current-voltage supralinearity, and open-channel noise through interactions with water or lipid headgroup atoms including the lipid ester carbonyl.
Seoh
S A
SA
Department of Physiology, Brown University, Providence, Rhode Island 02912, USA.
Busath
D
D
D000578:Amidines; D004594:Electrophysiology / Q000379:methods; D006096:Gramicidin; D007473:Ion Channels; D007700:Kinetics; D008051:Lipid Bilayers; D046911:Macromolecular Substances; D008564:Membrane Potentials; D008954:Models, Biological; D010649:Phenylalanine; D013329:Structure-Activity Relationship; D014364:Tryptophan
D000578:Amidines; D007473:Ion Channels; D008051:Lipid Bilayers; D046911:Macromolecular Substances; D006096:Gramicidin; C077922:formamidine; D010649:Phenylalanine; D014364:Tryptophan