Physics Colloquium
Friday, April 14th, 2004 2 P.M.

Prithwish Pal

University of Rochester Medical Center

'Illuminating a membrane transporter' - Resonance Energy Transfer as a tool for investigating structure and function of Anion Exchanger1

The human erythrocyte anion exchanger 1 (hAE1) is a polytopic membrane protein that has two distinct structural and functional domains. The N-terminal cytosplasmic domain (cdAE1) serves as an anchorage for red blood cell cytoskeleton and other cytosplasmic proteins such as hemoglobin. The cdAE1 undergoes a pH dependant conformational change that could be important in regulating the binding of cytosplasmic proteins. The C-terminal membrane embedded domain (mdAE1) is necessary and sufficient for an obligatory one for one exchange of chloride and bicarbonate across the red cell membrane, which plays a role in increasing the carbon dioxide carrying capacity of blood. The mdAE1 exists in two conformational states as it goes through the transport cycle, the inward facing form where it can bind anions only from the cytoplasm and the outward facing where it is accessible to anions only from the outside. Interconversions between the states occur only in the substrate bound form and understanding the differences in the conformation can give us important clues about how membrane transporters function. We have probed these different conformational states of mdAE1 as well as the cdAE1 using fluorescence tools such as steady-state and time-resolved fluorescence resonance energy transfer (FRET), time-resolved lanthanide energy transfer (LRET), and most recently with single pair FRET (spFRET).