Benzodiazepine-Mediated Structural Changes in the Multidrug Transporter P-Glycoprotein: An Intrinsic Fluorescence Quenching Analysis

• Type: Article in International Scientific Journal
• Date: 2008

Authors

Sofia A. C. Lima (Author) Other; Anabela Cordeiro-da-Silva (Author) FFUP; de Castro, B (Author) FCUP; Paula Gameiro (Author) Other

Journal

Title: Journal of Membrane Biology

Vol. 223 No. 3

Pages: 117-125

ISSN: 0022-2631

Publisher: Springer Nature

Indexing

ISI Web of Knowledge - 2 Citations

ISI Web of Science

Scopus - 3 Citations

Scientific classification

FOS: Medical and Health sciences > Other medical sciences

CORDIS: Health sciences

Other information

Authenticus ID: P-003-Z20

Resumo (PT): P-glycoprotein expressed in Pichia pastoris was used to study the drug binding sites of different benzodiazepines. The effect of bromazepam, chlordiazepoxide, diazepam and flurazepam on P-glycoprotein structure was investigated by measuring the intrinsic fluorescence of the transporter tryptophan residues. Purified mouse mdr1a transporter in mixed micelles of 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonic acid and 1,2-dimiristoyl-sn-glycerol-3-phosphocholine emitted fluorescence at 340 nm indicative of the fluorophores in a relatively apolar environment. Acrylamide and iodide ion were used as collisional quenchers toward distinct regions of the transporter, the protein and the interface protein-surface, respectively. Binding of ATP induced conformational changes at the protein surface level in accordance with the location of the nucleotide binding sites. Bromazepam interaction with the transporter was located at the protein-surface interface, diazepam at the membrane region and chlordiazepoxide at the protein surface. Only the flurazepam interaction site was not detected by the quenchers used. All benzodiazepines were able to elicit reorientation of the protein fluorophores on the P-glycoprotein—ATP complex. <br> <br> Keywords: P-glycoprotein - Intrinsic fluorescence - Benzodiazepine - ATP - Acrylamide quenching - Iodide ion quenching <br> <a target="_blank" href="http://www.springerlink.com/content/b57hjn483838680l/?p=f360a3a74e484aaca21bd2f1ec2a9c2a&pi=0"> Texto integral</a> <br> <br>

Abstract (EN): P-glycoprotein expressed in Pichia pastoris was used to study the drug binding sites of different benzodiazepines. The effect of bromazepam, chlordiazepoxide, diazepam and flurazepam on P-glycoprotein structure was investigated by measuring the intrinsic fluorescence of the transporter tryptophan residues. Purified mouse mdr1a transporter in mixed micelles of 3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonic acid and 1,2-dimiristoyl-sn-glycerol-3-phosphocholine emitted fluorescence at 340 nm indicative of the fluorophores in a relatively apolar environment. Acrylamide and iodide ion were used as collisional quenchers toward distinct regions of the transporter, the protein and the interface protein-surface, respectively. Binding of ATP induced conformational changes at the protein surface level in accordance with the location of the nucleotide binding sites. Bromazepam interaction with the transporter was located at the protein-surface interface, diazepam at the membrane region and chlordiazepoxide at the protein surface. Only the flurazepam interaction site was not detected by the quenchers used. All benzodiazepines were able to elicit reorientation of the protein fluorophores on the P-glycoprotein—ATP complex. <br> <br> Keywords: P-glycoprotein - Intrinsic fluorescence - Benzodiazepine - ATP - Acrylamide quenching - Iodide ion quenching <br> <a target="_blank" href="http://www.springerlink.com/content/b57hjn483838680l/?p=f360a3a74e484aaca21bd2f1ec2a9c2a&pi=0"> Full text</a> <br> <br>

Language: Portuguese

Type (Professor's evaluation): Scientific