Characterization of CO2/carbonic acid mediated proton flux through phosphatidylcholine vesicles as model membranes.

The apparent proton permeability coefficient for phospholipid vesicles measured in our laboratory (Norris, F. A. and Powell, G. L. (1990) Biochim. Biophys. Acta 1030, 165-171) for proton flux initiated by rapidly lowering of the external pH (acid jump) was a linear function of the reciprocal internal proton concentration. This behavior was ascribed to the presence of the weak acid carriers, carbonic acid/CO2/bicarbonate. In the present work, a theoretical description, appropriate for proton transport by any weak acid carrier, has been developed which lends itself to novel graphical treatment permitting the separate estimation of the permeability coefficients for protons, hydroxide ions and bicarbonate. The proton permeability coefficient determined by this method was 1.8 x 10(-5) (S.E. 1.3 x 10(-5)) cm/s; that for hydroxide ion was 3.8 x 10(-5) (S.E. 5.6 x 10(-6)) cm/s and a lower limit for the permeability of bicarbonate ion, 4.3 x 10(-6) (S.E. 3.6 x 10(-7) cm/s, can be set. The presence of negative surface charge on the lipid bilayer increased the observed proton permeability coefficient in accordance with Gouy-Chapman theory. The charge was introduced by preparing vesicles containing increasing amounts of negatively charged dioleoylphosphatidylglycerol. The observed proton permeability coefficient increased and the observed permeability coefficients for hydroxide ion and bicarbonate decreased. The addition of the lipophilic cations, valinomycin-K+ and tetrabutylammonium ion increased the slope of P vs. 1/[Hi+]. These changes are analogous to those reported for the permeant weak acid uncouplers FCCP and CCCP. These studies demonstrated that CO2/carbonic acid was an effective carrier of protons across phospholipid model membranes.