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In position 7 l- and d-Tic-substituted oxytocin and deamino oxytocin: NMR study and conformational insights

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Abstract

Incorporation of l- or d-Tic into position 7 of oxytocin (OT) and its deamino analogue ([Mpa1]OT) resulted in four analogues, [l-Tic7]OT (1), [d-Tic7]OT (2), [Mpa1,l-Tic7]OT (3) and [Mpa1,d-Tic7]OT (4). Their biological properties were described by Fragiadaki et al. (Eur J Med Chem 42:799–806, 2007). Their NMR study (NOESY, TOCSY, 1H–13C HSQC spectra) is presented here. Analogues 1, 3 and 4 showed partial agonistic activity, analogue 2 was pure antagonist, suggesting that a cis conformation between residues 6 and 7 of the molecule does not result in antagonistic activity. However, the reduction in agonistic activity of analogues 1, 3 and 4 in comparison to oxytocin is consistent with the reduction of the trans conformation form. Binding affinity for the human oxytocin receptor with IC50 value of 130, 730, 103, and 380 nM for peptides 1, 2, 3, and 4, respectively, showed lower affinity in the case of d analogues. Deamination slightly increased the affinity. The existence of both cis and trans configurations of the Cys6-d-Tic7 bond is supported by observation of two sets of cross-peaks for 1H and 13C nuclei for most of the residues of the peptide not only in NOESY and TOCSY but also in 1H–13C HSQC spectra. The MS and HPLC indicate the presence of a single molecule/peptide, and NMR data thus suggest that this second set of peaks is due to the cis conformation.

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Abbreviations

OT:

Oxytocin

Mpa:

β-Mercaptopropionic acid

[Mpa1]OT, dOT:

Deamino-oxytocin

Tic:

1,2,3,4-Tetrahydroisoquinoline-3-carboxylic acid

Fmoc:

9-Fluorenylmethoxycarbonyl

But :

t-Butyl

Trt:

Trityl

DIC:

Diisopropylcarbodiimide

HOBt:

1-Hydroxybenzotriazole

DMF:

Dimethylformamide

DMSO:

Dimethylsulfoxide

TFA:

Trifluoroacetic acid

HPLC:

High-performance liquid chromatography

ESI-MS:

Electrospray ionization-Mass spectrometry

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Acknowledgments

Z.S. and G.A.S. wish to acknowledge EU-NMR program—Contract # RII3-026145 (CERM; Center of Magnetic Resonance, University of Florence) for access to NMR instrumentation. The work was partly supported by research project No. Z40550506 of the IOCBof the Academy of Sciences of the Czech Republic.

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Authors and Affiliations

  1. Department of Pharmacy, University of Patras, 26500, Patras, Greece

    Zinovia Spyranti, Maria Fragiadaki, Vassiliki Magafa, Georgios A. Spyroulias & Paul Cordopatis

  2. Department of Antimicrobial Peptides, Institute of Organic Chemistry and Biochemistry, Academy of Sciences of Czech Republic, Flemingovo square 2, 16610, Prague 6, Czech Republic

    Lenka Borovickova & Jirina Slaninova

  3. Department of Pharmacy, University of Patras, Panepistimioupoli-Rion, 26504, Patras, Greece

    Georgios A. Spyroulias

Authors
  1. Zinovia Spyranti
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  2. Maria Fragiadaki
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  3. Vassiliki Magafa
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  4. Lenka Borovickova
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  5. Georgios A. Spyroulias
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  6. Paul Cordopatis
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  7. Jirina Slaninova
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Corresponding author

Correspondence to Georgios A. Spyroulias.

Additional information

Abbreviations of common amino acids are in accordance with the recommendations of IUPAC-IUB Joint Commission on Biochemical Nomenclature: Arch Biochem Biophys 206 (1988) v–xxii, J Biol Chem 264 (1989) 668–673 and J Peptide Sci 12 (2006) 1–8.

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Spyranti, Z., Fragiadaki, M., Magafa, V. et al. In position 7 l- and d-Tic-substituted oxytocin and deamino oxytocin: NMR study and conformational insights. Amino Acids 39, 539–548 (2010). https://doi.org/10.1007/s00726-009-0470-1

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