Investigating synthetic P-regions from voltage-gated sodium channel at the conformational and functional levels.
Four peptides mimicking the four P-regions of the electric eel sodium channel were chemically synthesized to characterize their secondary structure and their contribution to the channel selectivity. Circular dichroism spectra of these peptides in trifluoroethanol demonstrate an important beta-sheet conformational component. This beta-sheet content is much enhanced upon interaction with phosphatidylcholine small unilamellar vesicles. As expected (and except for P of domain III), no significant voltage-dependence is revealed in either macroscopic or single-channel conductance experiments. The concentrations-dependences of macroscopic conductances suggest that tetramers are the membrane conducting aggregates. In asymmetric ionic conditions, these channels made up of P-peptides were mostly specific for sodium over chloride whilst caesium was largely excluded. Single-channel conductance analysis discloses a moderate selectivity for sodium over potassium for PI and PII. This selectivity is larger with PIII but inverted for PIV. Finally, a control random peptide of the same length and with a comparable mean hydrophibicity was also tested. Its conformation in TFE is mainly unordered and no activity was detected in planar lipid bilayers. The data suggest that the presumed selectivity filter may not assume a circular symmetry and that molecular recognition between the different P-regions has to be taken into account.
Cosette
P
P
URA 500 CNRS-Université de Rouen, Mont-Saint-Aignan, France.
Brachais
L
L
Bernardi
E
E
Duclohier
H
H
D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004593:Electrophorus; D015640:Ion Channel Gating; D008051:Lipid Bilayers; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D015222:Sodium Channels / Q000737:chemistry* / Q000502:physiology*
D008051:Lipid Bilayers; D010446:Peptide Fragments; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D015222:Sodium Channels; C020888:1,2-dielaidoylphosphatidylethanolamine; C028694:1-palmitoyl-2-oleoylphosphatidylcholine