Hydrogen-deuterium-exchange kinetics of the amide protons of oxytocin studied by nuclear magnetic resonance

Abstract

The contribution of intramolecular hydrogen bonding to the solution structure of oxytocin was evaluated by study of amide hydrogen exchange rates in D2O by Fourier transform 1H NMR spectroscopy. Resolution enhancement filtering was employed in the determination of individual pseudo-first-order rate constants. Apparent barriers to exchange of 0.5 and 0.6 kcal mol-1 were measured for Asn5 and Cys6 peptide NH, respectively. The slowing is best explained by steric hindrance to solvent access in the case of Asn5, while for the Cys6 participation in a weak intramolecular hydrogen bond is possible. Fourfold acceleration of base-catalyzed exchange was observed for Tyr2 NH; it is proposed that this is the result of electronic effects induced by hydrogen bonding of Cys1 C=0, either to Cys6 NH or to the N-terminal amino group. Exchange proceeds near the random coil limit for each of the remaining residues. Comparison with exchange data for the model tripeptide N-acetyl-L-prolyl-L-leucylglycinamide demonstrates no evidence of noncovalent association of the tocin ring with the tripeptide tail of the hormone.

Binding of oxytocin and 8-arginine-vasopressin to neurophysin studied by nitrogen-15 NMR using magnetization transfer and indirect detection via protons

“…Thus, any hydrogen bonding of this -NH to the protein is likely to be weak [cf., Witanowski et al (1977) and Pardi et al (1983)]. It is also possible that the change in proton exchange rate at this position reflects the formation in the bound state of a salt bridge between the hormone a-NH3+ and a protein carboxyl [e.g., Breslow (1984)]; the a-NH3+ is a potential contributor to the rate of exchange of the -NH proton at Tyr-2 (Krauss & Cowburn, 1981).…”