Effect of salts on the structure of a potent analog of growth hormone releasing hormone as determined by optical spectroscopy.
Optical spectroscopic methods (circular dichroism, analytical ultracentrifugation, and static light scattering) were employed to study the solution behavior of an N-terminal-acylated 76-residue analog of growth hormone releasing hormone (GHRH). The GHRH analog had a 30% helical configuration in aqueous acidic solution, unlike other GHRH analogs that had a random coil configuration in aqueous solutions, and self-associated. High concentrations (7 M) of urea were required to obtain monomeric peptide, but such urea concentrations unfolded the peptide. Therefore, the folding and self-association were related events. The self-association and helix content were increased by the addition of various cations (e.g., Na+, Ca2+). In 7 M urea, when these cations were added, the peptide started to refold toward its aqueous conformation in a pH dependent manner; at low pH (2.5-3.5) the peptide folded to approximately 50% of the native configuration. At neutral pH (> 6) only small changes were seen when salts were added. When CaCl2 was added to a solution containing 7 M urea at pH 2.5, the self-association of the peptide increased with the concentration of CaCl2. Therefore, the salt dependent self-association observed in aqueous solvents was present in 7 M urea. Residues 45-76 in this GHRH analog, which are not present outside the hypothalamus, are probably needed to interact with a folding chaperone in vivo and provide stability for successful membrane transport and maintenance of biological activity.
Needham
G F
GF
Eli Lilly and Company, Greenfield, IN 46140, USA.
Pekar
A H
AH
Havel
H A
HA
D000595:Amino Acid Sequence; D002942:Circular Dichroism; D004926:Escherichia coli / Q000378:metabolism; D013007:Growth Hormone-Releasing Hormone / Q000031:analogs & derivatives* / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008027:Light; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000096:biosynthesis / Q000737:chemistry; D011487:Protein Conformation; D011489:Protein Denaturation; D012492:Salts; D012542:Scattering, Radiation; D013056:Spectrophotometry, Ultraviolet; D013329:Structure-Activity Relationship; D014461:Ultracentrifugation
D010446:Peptide Fragments; D012492:Salts; D013007:Growth Hormone-Releasing Hormone