Transbilayer transport of phosphatidic acid in response to transmembrane pH gradients

Preliminary studies have shown that asymmetric transbilayer distributions of phosphatidic acid (PA) can be induced by transmembrane pH gradients (delta pH) in large unilamellar vesicles [Hope et al. (1989) Biochemistry 28, 4181-4187]. Here the mechanism of PA transport is examined employing TNS as a fluorescent probe of lipid asymmetry. It is shown that the kinetics of PA transport are consistent with the transport of the uncharged (protonated) form. Transport of the neutral form can be rapid, exhibiting half-times for transbilayer transport of approximately 25 s at 45 degrees C. It is also shown that PA transport is associated with a large activation energy (28 kcal/mol) similar to that observed for phosphatidylglycerol. The maximum induced transbilayer asymmetry of PA corresponded to approximately 95% on the inner monolayer for vesicles containing 5 mol % PA.