Emiko Okamura - Kinetics of Binding and Diffusivity of Leucine-Enkephalin in Large Unilamellar Vesicle by Pulsed-Field-Gradient 1H NMR in Situ

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      Publication Details (including relevant citation   information):

         N. Yoshii, T. Emoto, E. Okamura,Biophysics, 7, 105-111 (2011).    


      The kinetics of   binding, the diffusivity, and the binding   amount of a   neuropeptide, leucine-enkephalin (L-Enk) to   lipid bilayer membranes are quantified by   pulsed-field-gradient (PFG) 1H NMR in situ.   The peptide signal is analyzed by the solution of the Bloch   equation with exchange terms in the presence of   large unilamellar vesicles (LUVs) as   confined, but fluid model cell  membranes. Even in   the case that the membrane-bound and   the free states of L-Enk cannot be distinguished in the   one-dimensional NMR spectrum, the PFG technique unveils the bound   component of L-Enk after the preferential decay of the free   component at the high field gradient. In 100-nm   diameter LUVs consisting of egg phosphatidylcholine,   the rate constants   of the peptide binding and dissociation are 0.040  and   0.40  s-1 at   303 K. This means that the  lifetime  of the  peptide  binding is of the   order from  second  to   ten-second.The   diffusivity of the bound L-Enkis  5×10-12  m2/s, almost  60 times as   restricted as the movement of free L-Enk  at 303 K.  One-tenth of 5 mM   L-Enk is bound to 40 mM LUV. The binding free energy is   calculated to be -2.9 kJ/mol, the magnitude close to the thermal   fluctuation, 2.5 kJ/mol. The result   demonstrates the potential of PFG 1H NMR to quantify   molecular dynamics of the peptide binding to membranes.

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