2 MECHANISMS OF H+ OH- TRANSPORT ACROSS PHOSPHOLIPID VESICULAR MEMBRANE FACILITATED BY GRAMICIDIN-A/

Two rate-limiting mechanisms have been proposed to explain the gramici din channel facilitated decay of the pH difference across vesicular me mbrane (Delta pH) in the pH region 6-8 and salt (MCl, M(+) = K+, Na+) concentration range 50-300 mM. 1) At low pH conditions (similar to 6), H+ transport through the gramicidin channel predominantly limits the Delta pH decay rate. 2) At higher pH conditions (similar to 7.5), tran sport of a deprotonated species (but not through the channel) predomin antly limits the rate. The second mechanism has been suggested to be t he hydroxyl ion propogation through water chains across the bilayer by hydrogen bond exchange. In both mechanisms alkali metal ion transport providing the compensating flux takes place through the gramicidin ch annels. Such an identification has been made from a detailed study of the Delta pH decay rate as a function of 1) gramicidin concentration, 2) alkali metal ion concentration, 3) pH, 4) temperature, and 5) chang es in the membrane order (by adding small amounts of chloroform to ves icle solutions). The apparent activation energy associated with the se cond mechanism (similar to 3.2 kcal/mol) is smaller than that associat ed with the first mechanism (similar to 12 kcal/mol). In these experim ents, Delta pH was created by temperature jump, and vesicles were prep ared using soybean phospholipid or a mixture of 94% egg phosphatidylch oline and 6% phosphatidic acid.