ATP binding cassette (ABC) transporters regulate the lipid asymmetry of the plasma membrane by exporting a wide variety of lipids to the outer leaflet. P-glycoprotein (PGP) is an ABC transporter responsible for the efflux of a large number of hydrophobic small molecules, such as drugs and fluorescent dyes. The interactions of PGP with its local environment on the plasma membrane are studied here using confocal and total internal reflection fluorescence microscopy (TIRFM). A fusion protein of PGP with enhanced green fluorescent protein (EGFP) and several rhodamine-derivative and cyanine fluorescent dyes make the transporter and its substrates visible. Here we use fluorescence recovery after photobleaching (FRAP) techniques to assess bulk protein mobility on the membrane. To complement this technique, TIRFM is used to track single PGP molecules and PGP clusters across the surface of the plasma membrane. Using these microscopic methods, the localization and dynamics of PGP on the membrane in the inactive, active, and inhibited states are determined and a preliminary mechanistic model is proposed.

Membrane transport activity and dynamics of the ABC transporter, P-glycoprotein

242nd National Meeting (2011)

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