Binding of oxytocin and 8-arginine-vasopressin to neurophysin studied by /sup 15/N NMR using magnetization transfer and indirect detection via protons
NMR was used to monitor the binding to neurophysin of oxytocin and 8-arginine-vasopressin, /sup 15/N labeling being used to identify specific backbone /sup 15/N and /sup 1/H signals. The most significant effects of binding were large downfield shifts in the amino nitrogen resonance of Phe-3 of vasopressin and in its associated proton, providing evidence that the peptide bond between residues 2 and 3 of the hormones is hydrogen-bonded to the protein within hormone-neurophysin complexes. Suggestive evidence for hydrogen bonding of the amino nitrogen of Tyr-2 was also obtained in the form of decreased proton exchange rates on binding; however, the chemical shift changes of this nitrogen and its associated proton indicated that such hydrogen bonding, if present, is probably weak. Shifts in the amino nitrogen of Asn-5 and in the -NH protons of both Asn-5 and Cys-6 demonstrated that these residues are significantly perturbed by binding, suggesting conformational changes of the ring on binding and/or the presence of binding sites on the hormone outside the 1-3 region. No support was obtained for the thesis that there is a significant second binding site for vasopressin on each neutrophysin chain. The behavior of both oxytocin and vasopressin on binding was consistent with formation of 1:1 complexes in slow exchange with the free state under most pH conditions. At low pH there was evidence of an increased exchange rate. Additionally, broadening of /sup 15/N resonances in the bound state at low pH occurred without a corresponding change in the resonances of equilibrating free hormone. The results suggest significant conformational alteration in neurophysin-hormone complexes at low pH possibly associated with protonation of the carboxyl group of the hormone-protein salt bridge.
- Research Organization:
- Rockefeller Univ., New York, NY
- OSTI ID:
- 5400501
- Journal Information:
- Biochemistry; (United States), Vol. 26:20
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
OXYTOCIN
CONFIGURATION INTERACTION
NUCLEAR MAGNETIC RESONANCE
PROTEINS
VASOPRESSIN
ARGININE
CHEMICAL SHIFT
NITROGEN 15
NMR SPECTRA
OVERHAUSER EFFECT
PH VALUE
PROTONS
AMINO ACIDS
BARYONS
CARBOXYLIC ACIDS
ELEMENTARY PARTICLES
FERMIONS
HADRONS
HORMONES
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
NITROGEN ISOTOPES
NUCLEI
NUCLEONS
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
PEPTIDE HORMONES
PITUITARY HORMONES
RESONANCE
SPECTRA
STABLE ISOTOPES
550601* - Medicine- Unsealed Radionuclides in Diagnostics