Membrane position of a basic aromatic peptide that sequesters phosphatidylinositol 4,5 bisphosphate determined by site-directed spin labeling and high-resolution NMR.
The membrane interactions and position of a positively charged and highly aromatic peptide derived from a secretory carrier membrane protein (SCAMP) are examined using magnetic resonance spectroscopy and several biochemical methods. This peptide (SCAMP-E) is shown to bind to membranes containing phosphatidylinositol 4,5-bisphosphate, PI(4,5)P2, and sequester PI(4,5)P2 within the plane of the membrane. Site-directed spin labeling of the SCAMP-E peptide indicates that the position and structure of membrane bound SCAMP-E are not altered by the presence of PI(4,5)P2, and that the peptide backbone is positioned within the lipid interface below the level of the lipid phosphates. A second approach using high-resolution NMR was used to generate a model for SCAMP-E bound to bicelles. This approach combined oxygen enhancements of nuclear relaxation with a computational method to dock the SCAMP-E peptide at the lipid interface. The model for SCAMP generated by NMR is consistent with the results of site-directed spin labeling and places the peptide backbone in the bilayer interfacial region and the aromatic side chains within the lipid hydrocarbon region. The charged side chains of SCAMP-E lie well within the interface with two arginine residues lying deeper than a plane defined by the position of the lipid phosphates. These data suggest that SCAMP-E interacts with PI(4,5)P2 through an electrostatic mechanism that does not involve specific lipid-peptide contacts. This interaction may be facilitated by the position of the positively charged side chains on SCAMP-E within a low-dielectric region of the bilayer interface.
Ellena
Jeffrey F
JF
Department of Chemistry and Biophysics Program, and Department of Cell Biology, University of Virginia, Charlottesville, Virginia 22904-4319, USA.
Moulthrop
Jason
J
Wu
Jing
J
Rauch
Michelle
M
Jaysinghne
Sajith
S
Castle
J David
JD
Cafiso
David S
DS
D001665:Binding Sites; D006841:Hydrocarbons, Aromatic / Q000032:analysis / Q000737:chemistry; D008051:Lipid Bilayers / Q000032:analysis / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008565:Membrane Proteins / Q000032:analysis / Q000737:chemistry; D008567:Membranes, Artificial; D008956:Models, Chemical; D008958:Models, Molecular; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000032:analysis / Q000737:chemistry*; D010743:Phospholipids / Q000032:analysis / Q000737:chemistry*; D011485:Protein Binding; D013113:Spin Labels
D006841:Hydrocarbons, Aromatic; D008051:Lipid Bilayers; D008565:Membrane Proteins; D008567:Membranes, Artificial; D019269:Phosphatidylinositol 4,5-Diphosphate; D010743:Phospholipids; D013113:Spin Labels