Sav1866 from Staphylococcus aureus and P-glycoprotein: similarities and differences in ATPase activity assessed with detergents as allocrites.

The ATP-binding cassette exporters Sav1866 from Staphylococcus aureus and P-glycoprotein are known to share a certain sequence similarity and disposition for cationic allocrites. Conversely, the two ATPases react very differently to neutral detergents that have previously been shown to be inhibitory allocrites for P-glycoprotein. To gain insight into the functional differences of the two proteins, we compared their basal and detergent-stimulated ATPase activity. P-Glycoprotein was investigated in NIH-MDR1-G185 plasma membrane vesicles and Sav1866 in lipid vesicles exhibiting a membrane packing density and a surface potential similar to those of the plasma membrane vesicles. Under basal conditions, Sav1866 revealed a lower catalytic efficiency and concomitantly a more pronounced sodium chloride and pH dependence than P-glycoprotein. As expected, the cationic allocrites (alkyltrimethylammonium chlorides) induced similar bell-shaped activity curves as a function of concentration for both exporters, suggesting stimulation upon binding of the first and inhibition upon binding of the second allocrite molecule. However, the neutral allocrites (n-alkyl-β-d-maltosides and n-ethylene glycol monododecyl ethers) reduced P-glycoprotein's ATPase activity at concentrations well below their critical micelle concentration (CMC) but strongly enhanced Sav1866's ATPase activity even at concentrations above their CMC. The lack of ATPase inhibition at high concentrations of neutral of detergents could be explained by their comparatively low binding affinity for the transmembrane domains of Sav1866, which seems to prevent binding of a second inhibitory molecule. The high ATPase activity in the presence of hydrophobic, long chain detergents moreover revealed that Sav1866, despite its lower basal catalytic efficiency, is a more efficient floppase for lipidlike amphiphiles than P-glycoprotein.