1. A CD and an NMR study of multiple bradykinin conformations in aqueous trifluoroethanol solutions
J R Cann, X Liu, J M Stewart, L Gera, G Kotovych Biopolymers. 1994 Jul;34(7):869-78. doi: 10.1002/bip.360340706.
CD and nmr studies have been carried out on aqueous trifluoroethanol (TFE) solutions of bradykinin (BK) and a bradykinin antagonist. The CD results exhibit a striking effect of TFE on the spectra of BK, with sequence Arg-Pro-Pro-Gly-Phe-Ser-Pro-Phe-Arg, and the BK antagonist, with sequence D-Arg-Arg-Pro-Hyp-Gly-Thi-D-Ser-D-Cpg-Cpg-Arg [where Hyp is 4-hydroxy-L-proline; Thi refers to beta-(2-thienyl)-L-alanine and Cpg refers to alpha-cyclopentylglycine). The effect of increasing concentration of TFE in water on the difference ellipticity at 222 nm was examined and showed that BK may be a mixture of at least two different conformers, one of which largely forms when the TFE concentration is increased beyond 80%. The linear extrapolation of 100% of the difference ellipticity of BK at low TFE concentrations yields a value in agreement with that shown by the BK antagonist, indicating that the conformation of BK at the lower TFE concentrations is similar to that of the BK antagonist. The conformational analysis was carried out using both one-dimensional and two-dimensional 1H-nmr techniques. The total correlation spectroscopy (TOCSY) spectrum of BK in a 60/40% (v/v) TFE/H2O solution at 10 degrees C and a nuclear Overhauser effect spectroscopy (NOESY) spectrum that shows only sequential H alpha (i)-NH(i + 1) or the H alpha (i)-H delta delta' (i + 1) NOEs indicate that the majority of the molecules adopt an all-trans extended conformation. The TOCSY for BK in the 95/5% (v/v) TFE/H2O solution shows that there are two major conformations in the solution with about equal population. The NOESY experiment shows two new important cross peaks for one conformation, namely Pro2 (alpha)-Pro3 (alpha) and the Pro2 (alpha)-Gly4(NH), indicating a cis Pro2-Pro3 bond and a type VI beta-turn between residues Arg1 and Gly4 involving cis proline at position 3, respectively. The low temperature coefficient of Gly4 for this conformation suggests the presence of an intramolecular hydrogen bond, therefore a type VIa beta-turn is present. The other conformation is all trans and extended. The BK antagonist shows difference CD spectra in TFE solutions referred to H2O that are superficially indicative of a beta-bend.(ABSTRACT TRUNCATED AT 400 WORDS)
2. Synthesis and some pharmacological properties (4-beta-(2-thienyl)-L-alanine)oxytocin
C W Smith, G Skala, R Walter J Med Chem. 1978 Jan;21(1):115-7. doi: 10.1021/jm00199a022.
The synthesis and some biological activities of [4-beta-(2-thienyl)-L-alanine]oxytocin are reported. This analogue has been studied in an ongoing exploration of the biological effects of introducing amino acid residues with bulky hydrophobic side chains into the second corner position of the beta turn present in the conformation of the 20-membered ring portion of oxytocin. The analogue was synthesized in stepwise manner by solution techniques utilizing ethylcarbamoyl protection for cysteine side chains. The presence of thienylalanine in position 4 evokes a drastic reduction in both affinity and intrinsic activity; the reduction in intrinsic activity was greater than that found for [Leu4]oxytocin or [Phe4]oxytocin. The analogue possesses 0.51 +/- 0.03 unit/mg of rat uterotonic potency and less than 0.05 unit/mg of rat pressor and rat antidiuretic potency and behaves as a competitive inhibitor of the response to oxytocin in the avian vasodepressor assay with a pA2 value of 7.44 +/- 0.19.
3. Synthesis and some pharmacological properties of (3-beta-(2-thienyl)-L-alanine)-8-lysine-vasopressin
C W Smith, M F Ferger, W Y Chan J Med Chem. 1975 Aug;18(8):822-5. doi: 10.1021/jm00242a012.
[3-beta-(2-Thienyl)-L-alanine]-8-lysine-vasopressin was synthesized by solution techniques. The partially protected heptapeptide Boc-Cys(Ec)-Tyr-Thi-Gln-Asn-Cys(Ec)-Pro (1) was synthesized in a stepwise manner using the active ester method or the dicyclohexylcarbodiimide (DCC) coupling technique mediated by 1-hydroxybenzotriazole (HBt). The protected nonapeptide amide Boc-Cys(Ec)-Tyr-Thi-Gin-Asn-Cys(Ec)-Pro-Lys(Coc)-Gly-NH2 (2) was prepared by coupling 1 with Lys(Coc)-Gly-NH2 using DCC-HBt. From 2, [3-thienylalanine]-8-lysine-vasopressin was obtained by removing the Boc-protecting groups with trifluoroacetic acid and ethylcarbamoyl (Ec) protecting groups in refluxing liquid NH3 followed by oxidative cyclization in H2O-MeOH using ICH2CH2I. Purification was effected by partition chromatography followed by gel filtration. The highly purified product possesses activities in the oxytocic, avian vasodepressor, rat pressor, and antidiuretic assays of 19.0 +/- 0.5, 87 +/- 4, 243 +/- 5, and 332 +/- 32 units/mg, respectively. Thus [3-thienylalanine]-8-lysine-vasopressin has higher oxytocic, avian vasodepressor, and antidiuretic potencies than does 8-lysine-vasopressin, whereas its pressor potency is about the same as or slightly lower than that of 8-lysine-vasopressin.