A fluorometric study of the interaction of bradykinin with lipids.
The interaction of bradykinin (BK) with lipids has been followed by steady-state fluorescence measurements. Addition of either cerebroside sulfate (CS) or phosphatidylinositol (PI), solubilized with the nonionic surfactant C12E8, to BK or its analogue [Gly6]-BK enhances the relative fluorescence intensity of peptide emission at 288 nm. Fluorometric titration of the peptide with lipid has been used to quantitate the interactions in terms of stoichiometry and equilibrium constant. Job's method of continuous variation for the BK-CS interaction gave a stoichiometry of 1:2 for the complex. The value of the equilibrium constant, K, for the interaction of either BK or [Gly6]-BK with CS is 1.5.10(4) M-1. The BK-PI interaction is weaker; K = 5.0.10(3) M-1. Although electrostatic forces no doubt play a major role in these interactions, measurements on the model peptide Gly-Phe-Gly indicate that the phenylalanine residues of BK are disposed in the hydrophobic environment provided by the lipid-C12E8 mixed micelle. 13C-NMR measurements on [99% 13C alpha-Gly6]-BK show that there is no change in its cis/trans ratio upon interaction with CS. The increase in the relative fluorescence intensity of BK accompanying its cooperative interaction with sodium dodecyl sulfate (SDS) implicates the role of hydrophobic forces in this interaction as well. These results bear on the interpretation of the changes in circular dichroism (CD) of BK caused by SDS.
Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver 80262.
A G
AG
Rao
J M
JM
Stewart
R J
RJ
Vavrek
L O
LO
Sillerud
N H
NH
Fink
J R
JR
Cann
D000595:Amino Acid Sequence; D001920:Bradykinin / Q000031:analogs & derivatives / Q000138:chemical synthesis; D002554:Cerebrosides; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols; D013050:Spectrometry, Fluorescence
D002554:Cerebrosides; D010716:Phosphatidylinositols; D001920:Bradykinin