title: Multiple ligand-binding properties of the lipocalin member chicken alpha1-acid glycoprotein studied by circular dichroism and electronic absorption spectroscopy: the essential role of the conserved tryptophan residue.
abstract: Multiple ligand-binding properties of the 30-kDa chicken alpha(1)-acid glycoprotein (cAGP), a member of the lipocalin protein family, were investigated for the first time by using circular dichroism (CD) and UV/Vis absorption spectroscopy methods. By measuring induced CD (ICD) spectra, high-affinity binding (K(a) approximately 10(5)-10(6) M(-1)) of several drugs, dyes and natural compounds to cAGP was demonstrated including antimalarial agents (quinacrine, primaquine), phenotiazines (chlorpromazine, methylene blue), propranolol, non-steroidal antiinflammatory drugs (ketoprofen, diclofenac), tamoxifen, diazepam, tacrine, dicoumarol, cationic dyes (auramine O, thioflavine T, ethidium bromide), benzo[a]pyrene, L-thyroxine, bile pigments (bilirubin, biliverdin), alkaloids (piperine, aristolochic acid), saturated and unsaturated fatty acids. Analysis of the extrinsic CD spectra with the study of the covalently modified protein and CD displacement experiments revealed that a single Trp26 residue of cAGP conserved in the whole lipocalin family is part of the binding site, and it is essentially involved in the ligand-binding process via pi-pi stacking interaction resulting in the appearance of strong induced CD bands due to the non-degenerate intermolecular exciton coupling between the pi-pi* transitions of the stacked indole ring-ligand chromophore. The finding that cAGP is able to accommodate a broad spectrum of ligands belonging to different chemical classes suggests that its core beta-barrel cavity is unusually wide containing overlapping sub-sites. Significance of these new data in understanding of the ligand-binding properties of other lipocalins, especially that of human AGP, and potential practical applications are briefly discussed. Overall, cAGP serves as a simple, ultimate model to extend our knowledge on ligand-binding properties of lipocalins and to study the role of tryptophan residues in molecular recognition processes.
authors.affiliation: Department of Molecular Pharmacology, Institute of Biomolecular Chemistry, Chemical Research Center, Budapest, P.O. Box 17, H-1525, Hungary. zsferi@chemres.hu
authors.forename: Ferenc
authors.initials: F
authors.lastname: Zsila
authors.forename: Hisami
authors.initials: H
authors.lastname: Matsunaga
authors.forename: Zsolt
authors.initials: Z
authors.lastname: Bikádi
authors.forename: Jun
authors.initials: J
authors.lastname: Haginaka
mesh_terms: D000818:Animals; D002645:Chickens; D002942:Circular Dichroism; D008024:Ligands; D009961:Orosomucoid / Q000737:chemistry / Q000378:metabolism*; D004364:Pharmaceutical Preparations / Q000378:metabolism; D013056:Spectrophotometry, Ultraviolet; D013057:Spectrum Analysis / Q000379:methods*; D014364:Tryptophan / Q000378:metabolism*
chemical_list: D008024:Ligands; D009961:Orosomucoid; D004364:Pharmaceutical Preparations; D014364:Tryptophan