Gramicidin structure and disposition in highly curved membranes.
With a view to deciphering aspects of the mechanism of membrane protein crystallization in lipidic mesophases (in meso crystallization), an examination of the structure and disposition of the pore-forming peptide, gramicidin, in the lipidic cubic phase was undertaken. At its simplest, the cubic phase consists of lipid and water in the form of a molecular 'sponge.' The lipid exists as a continuous, highly curved bilayer that divides the aqueous component into two interpenetrating but non-contacting channels. In this study, we show that gramicidin reconstitutes into the lipid bilayer of the cubic phase and that it adopts the channel, or helical dimer, conformation therein. Fluorescence quenching with brominated lipid was used to establish the bilayer location of the peptide. Electronic absorption and emission spectroscopies corroborated this finding. Peptide conformation in the cubic phase membrane was determined by circular dichroism. The identity and microstructure of the mesophases, and their capacity to accommodate gramicidin and other system components (sodium dodecyl sulfate, trifluoroethanol), was established by small-angle X-ray diffraction. Beyond a limiting concentration, gramicidin destabilized the cubic phase in favor of the inverted hexagonal phase. While gramicidin remained bilayer bound as membrane thickness changed, its conformation responded to the degree of bilayer mismatch with the hydrophobic surface of the peptide. These findings support the hypothesis that reconstitution into the lipid bilayer is an integral part of the in meso crystallization process as applied to membrane proteins. They also suggest ways for improving the process of membrane protein crystallogenesis.
Liu
W
W
The Ohio State University, Columbus, OH 43210, USA.
Caffrey
M
M
D000900:Anti-Bacterial Agents / Q000737:chemistry*; D006096:Gramicidin / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D008823:Micelles; D015394:Molecular Structure; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D014961:X-Ray Diffraction
D000900:Anti-Bacterial Agents; D008565:Membrane Proteins; D008823:Micelles; D012997:Solvents; D006096:Gramicidin; D012967:Sodium Dodecyl Sulfate