Conformational analysis of thyrotropin releasing factor by proton magnetic resonance spectroscopy

Abstract

SynopsisThe proton magnetic resonance spectrum of thyrotropin releasing factor (TRF) in solution in deuterium oxide and deuterated dimethylsulfoxide (DMSO-d6) has been analyzed. Two forms differing in cis-trans isomerism about the His-Pro peptide bond are observed. From the temperature dependence of chemical shift of the amide protons, it is concluded that TRF in DMSO-d6 does not contain intramolecular hydrogen bonds. Measurement of NH-C,H coupling constant provides an estimate of the histidine dihedral angle 4. Structural information about the histidine sidechain is deduced frcm C,H-CpH coupling constants and from the nonequivalence of the two prolyl &-protons. In DMSO-ds, there is evidence for a tautomeric equilibrium corresponding to an exchange of imidazole proton between the two nitrogen atoms N-6 and N-E. In water, the N-EH tautomer is found to be the predominant tautomeric form of the imidazole ring. These resultd in combination with energy calculation, vibrational analysis, and carbon nmr studies allow the determination of the conformation of TRF.

Ab-initio molecular orbital study of the cis/trans conformations of the peptide bond

“…The presence of cis peptide units has also been detected in small proline-containing, open-chain peptides, such as Gly-Pro-Leu [3] and the thyrotropin releasing factor pGlu-His-Pro-NH2 [4]. Despite the higher intrinsic energy of the cis peptide unit, the cis conformation has been proposed in residues of some fibrous proteins [ 5, 61. In view of the interest in the geometry of the peptide bond, many theoretical and experimental studies have been reported, especially on peptide analogs, such as N-monosubstituted amides.…”

The two conformations of TRH in solution

“…Whereas this case hardly presents a problem, the determination of the rotational angle 4'~ in histidine is more difficult as the corresponding coupling constant 3JcaH_NH = 7.5 Hz allows at least for three angular solutions: -150 °, -90 ° and +60 ° [9][10][11]. This ambiguity, due to the degeneracy of the Karplus curves was eliminated in this particular case with the help of the coupling constant 3JCrGlu_H~is (~ 2 Hz) obtained from compound I.…”

“…This was determined from the coupling constant 3JcaH_NH = 0 Hz [9][10][11]. Whereas this case hardly presents a problem, the determination of the rotational angle 4'~ in histidine is more difficult as the corresponding coupling constant 3JcaH_NH = 7.5 Hz allows at least for three angular solutions: -150 °, -90 ° and +60 ° [9][10][11].…”

Synthesis and conformational studies by 1H‐ and 13C‐nmr spectroscopy of a novel, sterically constrained analogue of thyrotropin‐releasing hormone

“…This indicates that, as in TRH,6~7~35-36 the pyroglutamyl residue is relatively free to rotate around t.he C"-C' bond A #, value of -20" was determined for the pyroglutamyl residue of TRH in water by Raman spectroscopy. 6 Rotations about both backbone torsional angles +2 and +? of the histidyl residue are allowed.…”