Nature of ligand affinity and dimerization of corticotrophin-releasing factor-binding protein may be detected by circular dichroism.
As the association of corticotrophin-releasing factor (CRF) with its binding protein (BP) to form a dimer complex (CRF2/BP2) appears to be dependent on the nature of the ligand we have compared the circular dichroism difference spectra after association of the BP with ovine (o) CRF, human (h) CRF and the alpha-helical CRF (9-41) antagonist. All three ligands caused a negative change in molar ellipticity above 210 nm, with oCRF having the least and hCRF the greatest effect. Below 210 nm there was a marked divergence of difference spectra, with the reaction with the natural peptides, hCRF and oCRF, resulting in a positive change in ellipticity, whilst that with the antagonist produced a negative change. In view of the BP spectrum indicating predominantly beta-sheet and the peptides showing mainly alpha-helix these results were interpreted as the changes above 210 nm being due to dimerization and below 210 nm to a change in the conformation of ligand on binding. The opposite change in alpha-helicity of the antagonist observed on binding compared with the two natural CRF peptides could have fundamental pharmacological implications.
Lowry
P J
PJ
School of Animal and Microbial Sciences, University of Reading, UK.
Koerber
S C
SC
Woods
R J
RJ
Baigent
S
S
Sutton
S
S
Behan
D P
DP
Vale
W
W
Rivier
J
J
D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D002850:Chromatography, Gel; D002942:Circular Dichroism; D003346:Corticotropin-Releasing Hormone / Q000737:chemistry / Q000378:metabolism*; D006727:Hormone Antagonists / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008024:Ligands; D046911:Macromolecular Substances; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012756:Sheep; D013056:Spectrophotometry, Ultraviolet
D002352:Carrier Proteins; D006727:Hormone Antagonists; D008024:Ligands; D046911:Macromolecular Substances; D010446:Peptide Fragments; D011994:Recombinant Proteins; C067133:corticotropin releasing factor-binding protein; D003346:Corticotropin-Releasing Hormone; C045089:corticotropin releasing hormone (9-41)