Cis→Trans Isomerization of Pro in Oxytocin Regulates Zn Binding.

Ion mobility/mass spectrometry techniques are employed to investigate the binding of Zn to the nine-residue peptide hormone oxytocin (OT, Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly-NH, having a disulfide bond between Cys and Cys residues). Zn binding to OT is known to increase the affinity of OT for its receptor [Pearlmutter, A. F., Soloff, M. S.: Characterization of the metal ion requirement for oxytocin-receptor interaction in rat mammary gland membranes. J. Biol. Chem. 254, 3899-3906 (1979)]. In the absence of Zn, we find evidence for two primary OT conformations, which arise because the Cys-Pro peptide bond exists in both the trans- and cis-configurations. Upon addition of Zn, we determine binding constants in water of K = 1.43 ± 0.24 and 0.42 ± 0.12 μM, for the trans- and cis-configured populations, respectively. The Zn bound form of OT, having a cross section of Ω = 235 Å, has Pro in the trans-configuration, which agrees with a prior report [Wyttenbach, T., Liu, D., Bowers, M. T.: Interactions of the hormone oxytocin with divalent metal ions. J. Am. Chem. Soc. 130, 5993-6000 (2008)], in which it was proposed that Zn binds to the peptide ring and is further coordinated by interaction of the C-terminal, Pro-Leu-Gly-NH, tail. The present work shows that the cis-configuration of OT isomerizes to the trans-configuration upon binding Zn. In this way, the proline residue regulates Zn binding to OT and, hence, is important in receptor binding. [ABSTRACT FROM AUTHOR]