Mapping the Binding Site of the Inhibitor Tariquidar That Stabilizes the First Transmembrane Domain of P-glycoprotein*
| Autor: | David M. Clarke, Tip W. Loo |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2015 |
| Předmět: |
Protein Folding
ATP Binding Cassette Transporter Subfamily B Glycoside Hydrolases Tariquidar ATP-binding cassette transporter Plasma protein binding Biology Arginine Biochemistry Protein Structure Secondary protein cross-linking Membrane Biology medicine Humans membrane protein Disulfides Binding site Molecular Biology P-glycoprotein Adenosine Triphosphatases Binding Sites Cell Membrane Cell Biology Transmembrane protein Drug Resistance Multiple Protein Structure Tertiary A-site Transmembrane domain Cross-Linking Reagents HEK293 Cells Mutation biology.protein Biophysics Quinolines ABC transporter Hydrophobic and Hydrophilic Interactions membrane enzyme medicine.drug Protein Binding |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 1083-351X 0021-9258 |
| Popis: | ABC (ATP-binding cassette) transporters are clinically important because drug pumps like P-glycoprotein (P-gp, ABCB1) confer multidrug resistance and mutant ABC proteins are responsible for many protein-folding diseases such as cystic fibrosis. Identification of the tariquidar-binding site has been the subject of intensive molecular modeling studies because it is the most potent inhibitor and corrector of P-gp. Tariquidar is a unique P-gp inhibitor because it locks the pump in a conformation that blocks drug efflux but activates ATPase activity. In silico docking studies have identified several potential tariquidar-binding sites. Here, we show through cross-linking studies that tariquidar most likely binds to sites within the transmembrane (TM) segments located in one wing or at the interface between the two wings (12 TM segments form 2 divergent wings). We then introduced arginine residues at all positions in the 12 TM segments (223 mutants) of P-gp. The rationale was that a charged residue in the drug-binding pocket would disrupt hydrophobic interaction with tariquidar and inhibit its ability to rescue processing mutants or stimulate ATPase activity. Arginines introduced at 30 positions significantly inhibited tariquidar rescue of a processing mutant and activation of ATPase activity. The results suggest that tariquidar binds to a site within the drug-binding pocket at the interface between the TM segments of both structural wings. Tariquidar differed from other drug substrates, however, as it stabilized the first TM domain. Stabilization of the first TM domain appears to be a key mechanism for high efficiency rescue of ABC processing mutants that cause disease. |
| Databáze: | OpenAIRE |
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