T A Scahill — Page 2 | PubMed Health

PubMedDate not supplied

1H NMR analysis and in vitro bioactivity of Leu27-bGRF(1-29)NH2 and its D-Ala2 and des-(Tyr1-Ala2)-analogs.

Relative growth hormone-releasing potencies of bovine growth hormone-releasing factor (bGRF) analogs bGRF(1-44)NH2 (I), Leu27-bGRF(1-29)NH2 (II) and D-Ala2, Leu27-bGRF(1-29)NH2 (III) in in vitro bovine anterior pituitary cell cultures were determined to be 100%, 48% and 77%, respectively. The potencies of II and III, although numerically different, were not statistically different. Leu27-bGRF(3-29)NH2 (IV) was approximately 10,000 times less potent than 1. 1H NMR studies of peptides II, III and IV in 35% d3-2,2,2-trifluorethanol (TFE)/65% phosphate buffer at pH 4 revealed very similar, highly helical secondary structures in the 8-29 region, with only subtle differences at the N-termini. This lack of correlation between secondary structure in solution and in vitro bioactivity suggests that either 1) the biological conformations induced at the GRF receptor for II and III vs. IV are different from those generated in TFE/buffer, 2) similar secondary structures may be necessary but not sufficient for the observed bioactivity or 3) residues 1 and 2 of analogs II and III are important contact residues crucial for effective GRF-receptor interaction.

Amino Acid Sequence↗

PubMedDate not supplied

Secondary structure of a human growth hormone-releasing factor fragment (Leu27-hGRF(15-32)NH2) in aqueous/SDS micelle environments.

The secondary structure of a human growth hormone-releasing factor (hGRF) fragment (Leu27-hGRF(15-32)NH2) has been studied by 1H NMR at 500 MHz in aqueous solutions containing varying concentrations of d25-sodium dodecyl sulfate (SDS). Chemical shifts, coupling constants and NOESY data show that the secondary structure of the peptide is random in aqueous solution in the absence of SDS. At relatively low molar ratios of SDS to peptide (1.3:1 to 3.3:1 SDS:peptide) the 1D 1H spectrum of the peptide changes as the peptide resonances are broadened significantly. NOESY patterns consistent with helical structure are present in the region of residues 22-29 when the SDS:peptide molar ratio is 1.3:1 and the SDS concentration is slightly below the critical micelle concentration (CMC). At higher molar ratios of SDS to peptide (16:1 to 72:1), where the SDS concentration is significantly above the CMC, the lineshape of the peptide's 1H NMR spectrum is sharpened. In these environments an alpha-helical conformation is induced in residues 19-32 of the hGRF fragment, as shown both by NOESY and by chemical shift data. Thus, the well-known tendency of this region of the GRF peptide to form alpha-helix in isotropic mixed-solvent systems (e.g., methanol/water, trifluoroethanol (TFE)/water) is seen also in SDS/aqueous systems.

Growth Hormone-Releasing Hormone↗