Sensitized photoinactivation of minigramicidin channels in bilayer lipid membranes.
The method of sensitized photoinactivation based on the photosensitized damage of gramicidin A (gA) molecules was applied here to study ionic channels formed by minigramicidin (the 11-residue analogue of gramicidin A) in a planar bilayer lipid membrane (BLM) of different thickness. Irradiation of BLM with a single flash of visible light in the presence of a photosensitizer (aluminum phthalocyanine or Rose Bengal) generating singlet oxygen provoked a decrease in the minigramicidin-induced electric current across BLM, the kinetics of which had the characteristic time of several seconds, as observed with gA. For gA, there is good correlation between the characteristic time of photoinactivation and the single-channel lifetime. In contrast to the covalent dimer of gA characterized by extremely long single-channel lifetime and the absence of current relaxation upon flash excitation, the covalent head-to-head dimer of minigramicidin displayed the flash-induced current decrease with the kinetics being strongly dependent on the membrane thickness. The current decrease became slower both upon increasing the concentration of the minigramicidin covalent dimer and upon including cholesterol in the membrane composition. These data in combination with the quadratic dependence of the current on the peptide concentration can be rationalized by hypothesizing that the macroscopic current across BLM measured at high concentrations of the peptide is provided by dimers of minigramicidin covalent dimers in the double beta(5.7)-helical conformation having the lifetime of about 0.4 s, while single channels with the lifetime of 0.01 s, observed at a very low peptide concentration, correspond to the single-stranded beta(6.3)-helical conformation. Alternatively the results can be explained by clustering of channels at high concentrations of the minigramicidin covalent dimer.
Dutseva
Elena A
EA
Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119992, Russia.
Antonenko
Yuri N
YN
Kotova
Elena A
EA
Pfeifer
Jochen R
JR
Koert
Ulrich
U
D019281:Dimerization; D004553:Electric Conductivity; D006096:Gramicidin / Q000378:metabolism*; D007473:Ion Channels / Q000378:metabolism* / Q000528:radiation effects*; D008027:Light; D008051:Lipid Bilayers / Q000378:metabolism*; D017319:Photosensitizing Agents / Q000494:pharmacology*; D013696:Temperature; D013997:Time Factors
D007473:Ion Channels; D008051:Lipid Bilayers; D017319:Photosensitizing Agents; D006096:Gramicidin