title: Determination of membrane Peptide orientation by 1H-detected 2H NMR spectroscopy.
abstract: We demonstrate the application of the proton inverse detected deuteron (PRIDE) NMR technique to the measurement of the orientation of membrane-bound peptides with enhanced sensitivity. Gramicidin D, a transmembrane peptide, and ovispirin, a surface-bound peptide, were used as model systems. The peptides were 2H-labeled by 1H/2H exchange and oriented uniaxially on glass plates. The directly detected 2H spectra of both peptides showed only a strong D(2)O signal and no large quadrupolar splittings. In contrast, the PRIDE spectrum of gramicidin exhibited quadrupolar splittings as large as 281 kHz, consistent with its transmembrane orientation. Moreover, the large D(2)O signal in the directly detected 2H spectra was cleanly suppressed in the PRIDE spectrum. For ovispirin, the 1H indirectly detected 2H spectrum revealed a 104 kHz splitting and a zero-frequency peak. The former reflects the in-plane orientation of most of the helix axis, while the latter results from residues with a magic-angle orientation of the N-D bonds. These are consistent with previous 15N NMR results on ovispirin. The combination of PRIDE and exchange labeling provides an economical and sensitive method of studying membrane peptide orientations in lipid bilayers without the influence of D(2)O and with the ability to detect N-D bonds at the magic angle from the bilayer normal.
authors.affiliation: Department of Chemistry, Iowa State University, Ames 50011, USA.
authors.forename: Satoru
authors.initials: S
authors.lastname: Yamaguchi
authors.forename: Mei
authors.initials: M
authors.lastname: Hong
mesh_terms: D023181:Antimicrobial Cationic Peptides / Q000737:chemistry*; D003903:Deuterium / Q000737:chemistry; D006096:Gramicidin / Q000737:chemistry*; D006859:Hydrogen / Q000737:chemistry; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008954:Models, Biological; D010455:Peptides / Q000737:chemistry*; D011487:Protein Conformation
chemical_list: D023181:Antimicrobial Cationic Peptides; D010455:Peptides; C440189:ovispirin; D006096:Gramicidin; D006859:Hydrogen; D003903:Deuterium