EVIDENCE FOR DIMER PARTICIPATION AND EVIDENCE AGAINST CHANNEL MECHANISM IN A23187-MEDIATED MONOVALENT METAL-ION TRANSPORT ACROSS PHOSPHOLIPID VESICULAR MEMBRANE

The decay of the pH difference (Delta pH) across soybean phospholipid vesicular membrane by ionophore A23187 (CAL)-mediated H+/M+ exchange ( M+ = Li+, Na+, K+, and Cs+) has been studied in the pH range 6-7.6. Th e Delta pH in these experiments were created by temperature jump. The observed dependence of Delta pH relaxation rate 1/tau on the concentra tion of GAL, pH, and the choice of M+ in vesicle solutions lead to the following conclusions. 1) The concentrations of dimers and other olig omers of A23187 in the membrane are small compared to the total concen tration of A23187 in the membrane, similar to that in chloroform solut ions reported in the literature. 2) In the H+ transport cycle leading to Delta pH decay, the A23187-mediated Hf translocation across the mem brane is a fast step, and the rate-limiting step is the A23187-mediate d M+ translocation. 3) Even though the monomeric Cal-H is the dominant species translocating H+, Cal-M is not the dominant species transloca ting M+ (even at concentrations higher than [Cal-H]), presumably becau se its dissociation rate is much higher than its translocation rate. 4 ) The pH dependence of 1/tau shows that the dimeric species Cal(2)LiLi , Cal(2)NaNa, Cal(2)KH, and Cal(2)CsH are the dominant species translo cating M+. The rate constant associated with their translocation has b een estimated to be similar to 5 x 10(3) s(-1). With this magnitude fo r the rate constants, the dimer dissociation constants of these specie s in the membrane have been estimated to be similar to 4, 1, 0.05, and 0.04 M, respectively. 5) Contrary to the claims made in the literatur e, the data obtained in the Delta pH decay studies do not favor the ch annel mechanism for the ion transport in this system. 6) However, they support the hypothesis that the dissociation of the divalent metal io n-A23187 complex is the rate limiting step of A23187-mediated divalent metal ion transport.