Dynamic properties of the backbone of an integral membrane polypeptide measured by 2H-NMR

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Abstract
The 2H-NMR spectrum of the exchangeable hydrogens of the synthetic amphiphilic polypeptide, lys2-gly-leu24-lys2-ala-amide, was measured for the solid peptide at room temperature and, as a function of temperature, for the peptide incorporated into hydrated dipalmitoylphosphatidylcholine (DPPC) bilayers. This study is a prototype of a similar class of experiments which can be carried out on integral membrane proteins to characterize, quantitatively, the dynamic properties of integral membrane proteins. At temperatures below the DPPC gel-liquid crystalline phase transition, the 2H NMR spectrum was very similar to that of the solid peptide indicating that the peptide was immobilized in the lipid bilayer on the time scale (≈10-5 s) of the 2H-NMR measurements. The 2H-NMR spectrum above the phase transition corresponded to that expected from a peptide in the α-helical conformation reorienting rapidly about the symmetry axis of the α-helix. Measurements of the quadrupolar echo relaxation time, T 2e , gave a quantitative measure of the correlation time, τ c , for this motion. The value of τ c decreased rapidly with increasing temperature as the fraction of DPPC molecules in the liquid crystalline phase increased, reaching a value of 2×10-7s above the phase transition. The observation of a characteristic minimum in T 2e as the temperature was raised provided a definitive, quantitative interpretation of the T 2e measurements. Using the known geometry of the peptide and the theory of uniaxial rotational diffusion, a value of η=1.1 poise was obtained for the effective viscosity of the membrane in close agreement with values obtained previously from transient linear dichroism measurements.