Photodynamic and radiolytic inactivation of ion channels formed by gramicidin A: oxidation and fragmentation.
Ion channels formed by the peptide gramicidin A in planar lipid membranes have been reported to react very sensitively upon irradiation of the membrane by ionizing radiation (radiolysis), by UV light (photolysis), or by visible light in the presence of appropriate photosensitizers (photodynamic inactivation). In all three cases the effect is due to the presence of the four tryptophan residues of the pentadecapeptide. Modifications of these amino acids--due to an interaction with free radicals formed upon water radiolysis or due to light absorption--have been found to reduce the membrane conductance by many orders of magnitude. The present study was intended to correlate functional changes, observed at the level of single ion channels, with changes of the molecular structure identified by mass spectrometry. About 98% of the inactivated channels showed a single-channel conductance of virtually zero, while about 2% of the channels present before irradiation are converted to a state of reduced conductance (and reduced lifetime). On the structural level, irradiation in the presence of the photosensitizer Rose Bengal was found to produce oxidation and fragmentation of the peptide at the positions of the tryptophan residues. Our results provide evidence that the main effect of radiolysis, or of photodynamic treatment, is the cleavage of the peptide backbone leading to immediate closure of an open ion channel.
Kunz
L
L
Department of Biology, University of Konstanz, Germany.
Zeidler
U
U
Haegele
K
K
Przybylski
M
M
Stark
G
G
D000595:Amino Acid Sequence; D004553:Electric Conductivity; D006096:Gramicidin / Q000737:chemistry / Q000528:radiation effects*; D066298:In Vitro Techniques; D007473:Ion Channels / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000528:radiation effects*; D007700:Kinetics; D008567:Membranes, Artificial; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010777:Photochemistry; D010782:Photolysis; D016339:Spectrometry, Mass, Fast Atom Bombardment; D014364:Tryptophan / Q000737:chemistry
D007473:Ion Channels; D008567:Membranes, Artificial; D006096:Gramicidin; D014364:Tryptophan