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NMR investigation of the electrostatic effect in binding of a neuropeptide, achatin-I, to phosphatidylcholine bilayers.

Kimura T, Ninomiya K, Futaki S

Institute for Chemical Research, Kyoto University, Uji, Kyoto 611-0011, Japan.

Achatin-I (Gly1-d-Phe2-Ala3-Asp4), known as a neuropeptide containing a d-amino acid, binds to the surface of a zwitterionic phosphatidylcholine (PC) membrane only when the peptide N-terminal amino group is in the ionized state, NH3+ (Kimura, T.; Okamura, E.; Matubayasi, N.; Asami, K.; Nakahara, M. Biophys. J. 2004, 87, 375-385). To gain mechanistic insights into how the binding equilibrium is delicately controlled by the ionization state of the N-terminal amino group, peptide-lipid binding interactions are investigated by selectively enriched 15N (at the N-terminus) and natural-abundance 13C NMR spectroscopy. Upon binding to the PC membrane, the 15N NMR of the N-terminal NH3(+) shifts upfield. This observation supports a mechanism that the role of the N-terminal NH3(+) in stabilizing the binding state is through electrostatic attraction with a headgroup negative charge, i.e., PO4(-). Interestingly, when the side chain beta-carboxyl group in Asp4 is deionized at acidic pH, the 15N signal of the N-terminal NH3(+) exhibits no significant chemical-shift change upon membrane binding of achatin-I. The Asp4 side chain thus regulates efficiency of the electrostatic binding between the peptide N-terminal NH3(+) and the lipid headgroup PO4(-). 13C chemical shifts in the hydrophobic D-Phe2 residue are largely perturbed upon membrane binding, in the case where the side chain beta-CO2(-) in Asp4 is deionized; the deionization of Asp4 beta-CO2(-) increases the net hydrophobicity of achatin-I with a reduction of both the electrostatic hydration and the electrostatic attraction with the headgroup N(CH3)3(+) in the most superficial region of the PC membrane, resulting in deeper anchoring of the phenyl ring. Hence, the electrostatic effect of the side chain beta-CO2(-) in Asp4 floats achatin-I on the PC membrane surface, and the binding equilibrium is sensitively controlled by the ionization state of the N-terminal NH3(+).

Published 5 April 2007 in J Phys Chem B, 111(14): 3831-8.
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