Induced chirality upon binding of cis-parinaric acid to bovine beta-lactoglobulin: spectroscopic characterization of the complex.
Binding of the polyunsaturated cis-parinaric acid to bovine beta-lactoglobulin (BLG) was studied by circular dichroism (CD), electronic absorption spectroscopy and mass spectrometry methods. Upon protein binding, the UV absorption band of parinaric acid is red shifted by ca. 5 nm, showing hypochromism and reduced vibrational fine structure, suggesting that the ligand binds as a monomer in non-planar geometry. In the CD spectra measured at pH 7.36 and 8.5 a strong, negative Cotton band appears centered at 310 nm (Delta epsilon = -25 M(-1) cm(-1)) corresponding to the long-wavelength absorption band of cis-parinaric acid. The source of this induced optical activity is the helical distortion of the polyene chromophore caused by the chiral protein environment. From CD spectral data the value of the association constant was calculated to be 4.7 x 10(5) M(-1) at pH 7.36. CD and mass spectrometry measurements showed that parinaric acid binds weakly to BLG in acidic solution, though small peaks at mass 18,559 and 18,645 can be obtained in the reconstructed electrospray mass spectrum; these correspond to the binding of parinaric acid in 1:1 stoichiometry to both monomer variants of BLG B and A. The hydrophobic interior cavity of BLG was assigned as the primary binding site of cis-parinaric acid.
Zsila
Ferenc
F
Department of Molecular Pharmacology, Institute of Chemistry, Chemical Research Center, P.O. Box 17, 1525 Budapest, Hungary. zsferi@chemres.hu
Imre
Tímea
T
Szabó
Pál T
PT
Bikádi
Zsolt
Z
Simonyi
Miklós
M
D000818:Animals; D002417:Cattle; D002942:Circular Dichroism; D005231:Fatty Acids, Unsaturated / Q000737:chemistry* / Q000378:metabolism; D007782:Lactoglobulins / Q000737:chemistry* / Q000378:metabolism; D013058:Mass Spectrometry / Q000379:methods; D008958:Models, Molecular; D011485:Protein Binding; D013056:Spectrophotometry, Ultraviolet / Q000379:methods
D005231:Fatty Acids, Unsaturated; D007782:Lactoglobulins; C013641:parinaric acid