Vinca alkaloid binding to P-glycoprotein occurs in a processive manner

Area of Science:

• Biochemistry
• Structural Biology
• Pharmacology

Background:

• P-glycoprotein (Pgp) transports a wide range of substrates, but its binding sites remain unclear.
• Previous studies suggested multiple binding sites, possibly in the central cavity or at the lipid-protein interface.

Purpose of the Study:

• To define the specific binding sites for vinblastine and rhodamine 123 on P-glycoprotein.
• To elucidate the mechanism of Pgp substrate binding and transport.

Main Methods:

• Site-directed mutagenesis of Pgp residues.
• Purification and reconstitution of Pgp into SMALPs and liposomes.
• Measurement of drug binding constants and ATP hydrolysis.
• Molecular docking of substrates to structural models of Pgp.

Main Results:

• Rhodamine 123 primarily binds within Pgp's central cavity.
• Vinblastine, a hydrophobic drug, binds at both the lipid-protein interface and the central cavity.
• A model was proposed where vinca alkaloids initially interact at the lipid interface before internalizing into the central cavity.

Conclusions:

• Pgp utilizes multiple initial interaction sites, including the lipid interface and central cavity, to accommodate diverse substrates.
• The central cavity serves as the transport conduit for Pgp substrates.
• This binding mechanism explains Pgp's broad substrate specificity.