title: Structural determinants for activation and block of CFTR-mediated chloride currents by apigenin.
abstract: Apigenin (4',5,7-trihydroxyflavone) is an activator of cystic fibrosis transmembrane conductance regulator (CFTR)-mediated Cl(-) currents across epithelia at low concentrations and a blocker at high concentrations. We determined the roles of structural components of apigenin for both stimulation and block of Cl(-) currents across Calu-3 epithelia. The half-maximal binding affinity of apigenin for current stimulation (K(s)) was 9.1 +/- 1.3 microM, and the rank-order of molecular structures was 7-hydroxyl > pyrone = 4'-hydroxyl > 5-hydroxyl. Both the 7-hydroxyl and the 4'-hydroxyl served as H-bond acceptors, whereas the 5-hydroxyl was an H-bond donor. The half-maximal binding affinity of apigenin during current block was 74 +/- 11 microM. Blocked Cl(-) currents were structurally determined by 7-hydroxyl = 4'-hydroxyl > pyrone > 5-hydroxyl. Prestimulation of tissues with forskolin significantly affected activation kinetics and binding characteristics. After forskolin stimulation, K(s) was 4.1 +/- 0.9 microM, which was structurally determined by pyrone > all hydroxyls > single hydroxyls. In contrast, block of Cl(-) current by apigenin was not affected by forskolin stimulation. We conclude that apigenin binds to a stimulatory and an inhibitory binding site, which are distinguished by their affinities and the molecular interactions during binding.
authors.affiliation: Children's Hospital Oakland Research Institute, Oakland 94609, California, USA.
authors.forename: B
authors.initials: B
authors.lastname: Illek
authors.forename: M E
authors.initials: ME
authors.lastname: Lizarzaburu
authors.forename: V
authors.initials: V
authors.lastname: Lee
authors.forename: M H
authors.initials: MH
authors.lastname: Nantz
authors.forename: M J
authors.initials: MJ
authors.lastname: Kurth
authors.forename: H
authors.initials: H
authors.lastname: Fischer
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000230:Adenocarcinoma; D000779:Anesthetics, Local / Q000494:pharmacology; D047310:Apigenin; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002712:Chlorides / Q000378:metabolism*; D005576:Colforsin / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000737:chemistry / Q000378:metabolism; D005419:Flavonoids / Q000737:chemistry / Q000494:pharmacology*; D006801:Humans; D008012:Lidocaine / Q000494:pharmacology; D008168:Lung / Q000166:cytology; D008175:Lung Neoplasms; D011802:Quinidine / Q000494:pharmacology; D000077185:Resveratrol; D013267:Stilbenes / Q000737:chemistry / Q000494:pharmacology; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured; D062367:ortho-Aminobenzoates / Q000494:pharmacology
chemical_list: D000779:Anesthetics, Local; C505032:CFTR protein, human; D002121:Calcium Channel Blockers; D002712:Chlorides; D004791:Enzyme Inhibitors; D005419:Flavonoids; D013267:Stilbenes; D062367:ortho-Aminobenzoates; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005576:Colforsin; D047310:Apigenin; C001209:fenamic acid; D008012:Lidocaine; D011802:Quinidine; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D000077185:Resveratrol