Small peptides as analogs of oxytocin and vasopressin in their interactions with bovine neurophysin-II

Abstract

Circular dichroism and proton titration studies of mixtures of native or nitrated bovine neurophysin-II and lysine vasopressin confirm that there is one principal site for lysine vasopressin of very similar properties to the single oxytocin site. A second, but markedly weaker lysine vasopressin site is allowed by the data, but two thermodynamically equivalent sites for lysine vasopressin are precluded. Binding constants to nitrated bovine neurophysin-II of oxytocin, lysine vasopressin, and a series of peptide analogs of the first two to three residues of the hormones were obtained by circular dichroism using a single-site model. The data indicate that peptides containing only the first three residues of the hormones contribute almost two-thirds of the binding free energy of the hormones and that half of the binding free energy is contributed by the cooperative binding of residues 1 and 2.

Crystal structure of a bovine neurophysin II dipeptide complex at 2.8 A determined from the single-wavelength anomalous scattering signal of an incorporated iodine atom.

“…The crystal structure also confirms the disulfide assignments of Burman et al (7) and is consistent with recent conclusions from solution data as to the relationship of specific protein residues to the principal hormone-binding site. The relationship ofTyr49 to the binding site has been particularly studied because of its marked perturbation by binding (8,38). In the crystal, the suggested proximity of this residue to the binding site (1,39,40) is confirmed, with the backbone atoms of Tyr49 approaching bound peptide at a distance slightly less than 6 A.…”

“…Bovine NP-I was purified as described (14). The dipeptide para-iodo-L-phenylalanyltyrosine amide (I-Phe-Tyr-NH2) was custom-synthesized by Peninsula Laboratories and was demonstrated by using circular dichroism (8) to bind to the hormone-binding site with high affinity. Crystals of the NP-dipeptide complex were grown at pH 7.5 by using a modification of the procedure of Yoo et al (9) Data Collection.…”

“…Solution studies have established the use of small di-and tripeptides analogs of the hormone amino termini as models for the hormones in their interactions with NP (8). We previously reported the crystallization ofa complex of bovine NP-II (a vasopressin-associated NP) with Phe-Tyr-NH2, which binds to the hormone-binding site analogously to the hormones.…”

“…We also crystallized des-(93-95)-NP-II as its Phe-Tyr-NH2 complex in three distinct crystal forms (10), one of which is isomorphous with the crystals of a porcine NP-I complex (11). These contained 4,8, and 12 molecules per asymmetric unit, respectively, suggesting that the basic aggregates of the NP-dipeptide complex in the crystals are tetramers that can self-associate to higher oligomers (10). In view of the probable importance of NP self-association in packaging of the precursor in the NSG (1,4) and evidence that NP complexes can be present in the NSG as the precipitated or crystalline state (12,13), analysis of the crystal structure of these complexes offers an opportunity for providing detailed structural information not only on NP folding and protein-peptide interactions but also on how the complexes might be packaged in NSG.…”

Structural basis of neurophysin hormone specificity: Geometry, polarity, and polarizability in aromatic ring interactions

“…The method used Breslow et al, 1973! is based on the large change in CD spectrum of the NP nitrotyrosine when the hormone-binding site is occupied.…”