Starting with assignments of proton nuclear magnetic resonance previously made for oxytocin in deuterated dimethylsulfoxide at 220 MHz, we have assigned resonances for the mammalian antidiuretic hormone, lysine vasopressin. The results demonstrate that spectral assignments of neurohypophyseal hormones and their congeners can, within certain limits, be derived from each other. Comparison of the spectra of lysine vasopressin and oxytocin suggests that the gross backbone conformations of their 20-membered ring components are for the most part similar in deuterated dimethylsulfoxide, whereas the C-terminal acyclic amino-acid sequence of lysine vasopressin is more flexible than that of oxytocin.

Proton nuclear magnetic resonances of two analogs of the mammalian antidiuretic hormone, lysine vasopressin, have been assigned. The analogs studied were deamino-lysine vasopressin and deamino-8-tosyllysine vasopressin, formally derived from lysine vasopressin by removal of the potential cationic sites. Most assignments made in deuterated dimethylsulfoxide at 220 MHz were directly derived from the already determined spectrum of lysine vasopressin in this solvent [Walter et al., Proc. Nat. Acad. Sci. USA (1972) 69, 1920]. Comparison of chemical shifts of peptide NH peaks, aCH-NH coupling constants, temperature dependence of peptide NH resonances, and proton-deuterium exchange rates revealed that from a conformational standpoint both deamino analogs occupy intermediary positions between lysine vasopressin and oxytocin.From the point of view of conformation, the most significant difference between the congeneric neurohypophyseal hormones, oxytocin and lysine vasopressin (LysVP), is the greater hydrophilicity of LysVP. In addition to the NH2-terminal amino group of Cys-1 that is present in both of these nonapeptide hormones, LysVP possesses a second potential cationic site in the e-amino group of the lysine residue in position 8. In order to evaluate the extent to which the presence of two amino groups in LysVP contributes to its conformational variances with respect to oxytocin (1-3), we have compared the proton magnetic properties of these two hormones with those of the two LysVP analogs, [1-f-mercaptopropionic acid, 8-lysine]-vasopressin (also referred to as deamino- Fig. 1), in which the number of primary amino groups has been reduced stepwise. In d-NH2LysVP the NHrterminal amino group is formally replaced by a hydrogen atom, and in dNH2TosLysVP an additional modification is introducedthe Camino group of Lys is tosylated. MATERIALS AND METHODSd-NH2LysVP and d-NH2TosLysVP were prepared by Havran (manuscript in preparation),-who used a solid-phase method of peptide synthesis (4, 5). This d-NH2LysVP exhibited a rat pressor activity (6) of 132 ± 7 U/mg, which is identical to the activity of 126 d-2 U/mg reported for this analog when it was prepared by conventional methods of peptide synthesis (7).Spectra were recorded on a Varian Associates HR-220 NMR Spectrometer, and proton double-resonance experiments were done as described (3) (3) Abbreviations: Me2SO, dimethylsulfoxide; Me4Si, tetramethylsilane; LysVP, lysine vasopressin;

The neurohypophyseal hormone [8-arginine]-vasopressin has been synthesized by the solid-phase method. The protected nonapeptide intermediate, S-benzyl-TV-tosyl-L-cysteinyl-L-tyrosyl-L-phenylalanyl-L-glutaminyl-L-asparaginyl-S-benzyl-L-cysteinyl-L-prolyl-TV°-tosyl-L-arginylglycinamide, has been obtained in crystalline form. Removal of the protecting groups from the crystalline protected nonapeptide followed by oxidative cyclization provided highly active hormone preparations in excellent yields. Further purification by ion exchange chromatography afforded [8-arginine]-vasopressin possessing 503 ± 53 units/per mg of rat antidiuretic activity, 487 ±15 units/mg of rat pressor activity, 12.0 ± 0.2 units/mg of rat oxytocic activity, 30-120 units/mg of rabbit milkejecting activity, and 100 ± 5 units/mg of avian vasodepressor activity.The solid-phase method of peptide synthesis6 hag been successfully used for the preparation of several naturally occurring neurohypophyseal peptides such as oxytocin,6 [8-lysine]-vasopressin,7 and glumitocin.8This paper describes the application of this method to the synthesis of the mammalian antidiuretic principle, [8-arginine]-vasopressin,9 Figure 1. Chiefly through the crystallization of the protected nonapeptide (S-benzyl-TV-tosyl-L-cysteinyl-L-tyrosyl-lphenylalanyl-L-glutaminyl-L-asparaginyl-S'-benzyl-Lcysteinyl -L-prolyl-TVG-tosyl-L-arginylglycinamide, I, Figure 2) the hormone was obtained in excellent yield and with high biological potency.Earlier syntheses of [8-arginine]-vasopressin gave preparations exhibiting considerable variation in the * To whom correspondence should be addressed.