13C-13C rotational resonance in a transmembrane peptide: a comparison of the fluid and gel phases.
A comparative study of two doubly 13C labeled amphiphilic transmembrane peptides was undertaken to determine the potential of rotational resonance for measuring internuclear distances through the direct dipolar coupling in the presence of motion. The two peptides, having the sequence acetyl-K2-G-L16-K2-A-amide, differed only in the position of 13C labels. The first peptide, [1-13C]leu(11):[alpha-13C]leu(12), had labels on adjacent residues, at the carbonyl of leu(11) and the alpha carbon of leu(12). The second, [1-13C]leu(8):[alpha-(13)/C]leu(11), was labeled on consecutive turns of the alpha-helical peptide. The internuclear distance between labeled positions of the first peptide, which for an ideal alpha helix has a value of 2.48 A, is relatively independent of internal flexibility or peptide conformational change. The dipolar coupling between these two nuclei is sensitive to motional averaging by molecular reorientation, however, making this peptide ideal for investigating these motions. The internuclear distance between labels on the second peptide has an expected static ideal alpha-helix value of 4.6 A, but this is sensitive to internal flexibility. In addition, the dipolar coupling between these two nuclei is much weaker because of their larger separation, making this peptide a much more difficult test of the rotational resonance technique. The dipolar couplings between the labeled nuclei of these two peptides were measured by rotational resonance in the dry peptide powders and in multilamellar dispersions with dimyristoylphosphatidylcholine in the gel phase, at -10 degrees C, and in the fluid phase, at 40 degrees C. The results for the peptide having adjacent labels can be readily interpreted in terms of a simple model for the peptide motion. The results for the second peptide show that, in the fluid phase, the motionally averaged dipolar coupling is too small to be measured by rotational resonance. Rotational resonance, rotational echo double resonance, and related techniques can be used to obtain reliable and valuable dipolar couplings in static solid and membrane systems. The interpretation of these couplings in terms of internuclear distances is straightforward in the absence of molecular motion. These techniques hold considerable promise for membrane protein structural studies under conditions, such as at low temperatures, where molecular motion does not modulate the dipolar couplings. However, a typical membrane at physiological temperatures exhibits complex molecular motions. In the absence of an accurate and detailed description of both internal and whole body molecular motions, it is unlikely that techniques of this type, which are based on extracting distances from direct internuclear dipolar couplings, can be used to study molecular structure under these conditions. Furthermore, the reduction in the strengths of the dipolar couplings by these motions dramatically reduces the useful range of distances which can be measured.
Langlais
D B
DB
Department of Physics, Simon Fraser University, Burnaby, British Columbia, Canada V5A 1S6.
Hodges
R S
RS
Davis
J H
JH
D000595:Amino Acid Sequence; D002247:Carbon Isotopes; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003460:Crystallization; D004134:Dimyristoylphosphatidylcholine / Q000737:chemistry; D005782:Gels; D008051:Lipid Bilayers / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry*; D008956:Models, Chemical; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry*; D013816:Thermodynamics
D002247:Carbon Isotopes; D005782:Gels; D008051:Lipid Bilayers; D008565:Membrane Proteins; D010455:Peptides; D004134:Dimyristoylphosphatidylcholine