Carbon-13 NMR studies of the Peptide hormones oxytocin, arginine vasopressin, isotocin, mesotocin, glumitocin, aspartocin, related analogs, and diastereoisomers. Use of specifically deuterated hormone derivatives for assignments and effects of structural changes on carbon-13 NMR chemical shifts in peptides

Hruby, Victor J.; Deb, K. K.; Spatola, Arno F.; Upson, Donald A.; Yamamoto, Diane M.

doi:10.1021/ja00495a033

Cited by 33 publications

(7 citation statements)

References 6 publications

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“…The proton decoupled 13 C‐NMR spectrum of the 13 C‐enriched OT compound 16 in D 2 O showed three signals at the expected chemical shifts for the corresponding carbons in OT: δ C=O = 167.7 ppm, δ Cα = 39.5 ppm, and δ Cβ = 52.2 ppm, J CN = 5.7 Hz . The proton decoupled 13 C‐NMR spectra of the dimeric degradation products 19a and 20a were obtained for both compounds and showed signals for six different carbon atoms (Table ).…”

Section: Resultsmentioning

confidence: 96%

On the mechanism of degradation of oxytocin and its analogues in aqueous solution

Wiśniewski¹,

Finnman²,

Flipo³

et al. 2013

Biopolymers

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Oxytocin (OT) is a cyclic nonapeptide containing one internal disulfide bond between its Cys(1) and Cys(6) residues. Although OT is one of the most commonly used peptidic drugs, the mechanism of its degradation in aqueous solution and the identity of its degradants have not been fully elucidated. To investigate the pathways and products of OT degradation in slightly acidic to neutral solutions, we prepared the peptides: OT, [D-Cys(1)]OT, a series of N-alkylated OT analogues, [[(13)C3,(15) N]Cys(1)]OT, and OT where each sulfur atom was systematically replaced by either methylene, (34)S, or Se. The peptides were incubated at 40°C and the degradation products studied by HPLC, LCMS, and (13)C-NMR. Our findings suggest that the degradation begins with β-elimination of the disulfide linkage to form a putative intermediate linear peptide containing an S-thiocysteine (a persulfide) in position 6 and a dehydroalanine in position 1. This intermediate persulfide appears to donate a sulfur atom to an intact OT molecule to form OT trisulfide and higher monomeric polysulfides, while the dehydroalanine residue is hydrolyzed with loss of the N-terminal amino group to yield a linear N-pyruvoylated octapeptide containing a reduced Cys(6). Based on the MS and (13)C-NMR data of the products from degradation of [[(13)C3,(15)N]Cys(1)]OT, we postulate that the ultimate degradation products of OT are dimers composed of two pyruvoylated octapeptides held together by one disulfide bridge between the two Cys(6) residues and by one more, non-reducible, linkage resulting from an aldol-type condensation between the two N-terminal pyruvoyl groups.

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Section: Resultsmentioning

confidence: 96%

On the mechanism of degradation of oxytocin and its analogues in aqueous solution

Wiśniewski¹,

Finnman²,

Flipo³

et al. 2013

Biopolymers

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“…This is a result of the multiple values of x* 1 for the tyrosine, the lack of information concerning the orientation of the tyrosine ring about the C^-C-y bond, and the lack of information concerning the lie3 x angles. Nonetheless, the lack of any significant chemical shift differences for the lie3 and Gin4 side chains in both the carbon-13 (Hruby et al, 1979a) and proton NMR spectra of [Pen1]oxytocin and [Pen*,Leu2]oxytocin suggests that a ring current effect is not a significant contributor to the observed differences.…”

Section: Discussionmentioning

confidence: 91%

“…troscopy (Meraldi et al, 1975(Meraldi et al, , 1977Hruby et al, 1979a), carbon-13 spin-lattice relaxation (7"j) studies (Meraldi et al, 1977) , and CD and laser Raman spectroscopy (Hruby et al, 1978) we showed that, while oxytocin had considerable conformational and dynamic flexibility, [Pen']oxytocin was conformationally and dynamically more restricted. The spectroscopic data were consistent with a conformation for the 20-membered ring moiety of [Pen*]oxytocin that possessed two l*-3 intramolecular hydrogen bonds (C7 structures) (Meraldi et al, 1975(Meraldi et al, , 1977Hruby, 1980), a disulfide bond with a dihedral angle of 110-115°and right-handed chirality (Hruby et al, 1978), and a restricted Tyr2 side chain in which the rotamer that would place the aromatic side chain pointing toward the 20-membered disulfide ring was excluded.…”

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confidence: 71%

Conformational study of the potent peptide hormone antagonist [1-penicillamine,2-leucine]oxytocin in aqueous solution

Mosberg¹,

Hruby²,

Meraldi³

1981

Biochemistry

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[1-Penicillamine,2-leucine]oxytocin is a conformationally restricted analogue of oxytoxin in which the half-cystine-1 and tyrosine-2 residues of the native hormone are replaced by half-penicillamine (beta, beta-dimethyl-half-cystine) and leucine, respectively. This analogue is a surprisingly potent oxytocin antagonist [Hruby, V. J., Deb, K. K., Yamamoto, D. M., Hadley, M. E., & Chan, W. Y. (1979) J. Med. Chem. 22,7]. Extensive proton magnetic resonance experiments were performed to determine the conformational properties of this analogue in aqueous solution, and the results were compared with the previously published model for the conformation of [1-penicillamine]oxytocin. The results are consistent with a conformation similar to that of [1-penicillamine]oxytocin except that, while [1-penicillamine]oxytocin in aqueous solution possesses two 1 comes from 3 (C7) type turns involving the isoleucine-3 peptide amide proton and the half-penicillamine-1 carbonyl and the asparagine-5 peptide amide proton and the isoleucine-3 carbonyl, [1-penicillamine,2-leucine)oxytocin has only the latter 1 comes from 3 turn. This difference between the antagonists is reflected in the different phi and psi angles in the three N-terminal residues of the two inhibitor analogues and in differences in the preferred side-chain conformations for several residues. One particular result of these conformational differences is that, whereas for [1-penicillamine]oxytocin the tyrosine-2 side chain is unable to assume the rotamer for maximal binding to the uterine receptor, [1-penicillamine,2-leucine]oxytoxin retains conformational and dynamic properties at residues two and three which are more similar to those of oxytocin. It is postulated that these conformational and dynamic properties are consistent with the stronger binding and, hence, greater antagonist activity for this penicillamine analogue relative to [1-penicillamine]oxytocin.

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“…Ironically, the revised structure for amphomycin is identical to one of the two possible structures originally proposed by Fujino in 1965 for glumamycin 22. Additionally, the term aspartocin has been used for an oxytocin‐type hormone with elemental formula C 42 H 64 N 12 O 12 S 2 23, 24 and its use should be avoided.…”

Section: Discussionmentioning

confidence: 94%

Structure characterization of lipocyclopeptide antibiotics, aspartocins A, B & C, by ESI‐MSMS and ESI‐nozzle‐skimmer‐MSMS

Siegel¹,

Kong²,

Feng³

et al. 2009

J. Mass Spectrom.

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Three lipocyclopeptide antibiotics, aspartocins A (1), B (2), and C (3), were obtained from the aspartocin complex by HPLC separation methodology. Their structures were elucidated using previously published chemical degradation results coupled with spectroscopic studies including ESI-MS, ESI-Nozzle Skimmer-MSMS and NMR. All three aspartocin compounds share the same cyclic decapeptide core of cyclo [Dab2 (Asp1-FA)-Pip3-MeAsp4-Asp5-Gly6-Asp7-Gly8-Dab9-Val10-Pro11]. They differ only in the fatty acid side chain moiety (FA) corresponding to (Z)-13-methyltetradec-3-ene-carbonyl, (+,Z)-12-methyltetradec-3-ene-carbonyl and (Z)-12-methyltridec-3-ene-carbonyl for aspartocins A (1), B (2), and C (3), respectively. All of the sequence ions were observed by ESI-MSMS of the doubly charged parent ions. However, a number of the sequence ions observed were of low abundance. To fully sequence the lipocyclopeptide antibiotic structures, these low abundance sequence ions together with complementary sequence ions were confirmed by ESI-Nozzle-Skimmer-MSMS of the singly charged linear peptide parent fragment ions H-Asp5-Gly6-Asp7-Gly8-Dab9-Val10-Pro11-Dab2(1+)-Asp1-FA. Cyclization of the aspartocins was demonstrated to occur via the beta-amino group of Dab2 from ions of moderate intensity in the ESI-MSMS spectra. As the fatty acid moieties do not undergo internal fragmentations under the experimental ESI mass spectral conditions used, the 14 Da mass difference between the fatty acid moieties of aspartocins A (1) and B (2) versus aspartocin C (3) was used as an internal mass tag to differentiate fragment ions containing fatty acid moieties and those not containing the fatty acid moieties. The most numerous and abundant fragment ions observed in the tandem mass spectra are due to the cleavage of the tertiary nitrogen amide of the pipecolic acid residue-3 (16 fragment ions) and the proline residue-11 (7 fragment ions). In addition, the neutral loss of ethanimine from alpha,beta-diaminobutyric acid residue 9 was observed for the parent molecular ion and for 7 fragment ions.

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Cited by 33 publications

References 6 publications

On the mechanism of degradation of oxytocin and its analogues in aqueous solution

On the mechanism of degradation of oxytocin and its analogues in aqueous solution

Conformational study of the potent peptide hormone antagonist [1-penicillamine,2-leucine]oxytocin in aqueous solution

Structure characterization of lipocyclopeptide antibiotics, aspartocins A, B & C, by ESI‐MSMS and ESI‐nozzle‐skimmer‐MSMS

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