Sizing of an ion pore by access resistance measurements.

Dr. Bezrukov is on leave from Leningrad Nuclear Physics Institute of Academy of Sciences of the Union of Soviet Socialist Republics, 188350 USSR. Indeed, the total resistance of the channel contains information about pore dimensions, but a direct evaluation of pore radius from R, is complicated by the uncertainty in the value of Rch' Unfortunately, the mechanisms of an ion transport within the ion channel are still poorly understood and theoretical evaluation of Reh even in the detailed structural models is quite problematic. Using water-soluble polymers of different molecular weight and physical properties we were able to separate contributions from the channel proper and the access resistance into the total resistance of the channel. In principle, addition of water-soluble polymers can change the total channel resistance due to following reasons. First, polymers of sufficiently small size can penetrate the channel, "block" the current paths, and increase the channel resistance proportionally to the polymer partition into the channel. Second, the polymer addition can influence dissolved salt activity, thus changing salt concentration inside the channel and, therefore, resistance of the channel proper. Third, a nonelectrogenic polymer increases the bulk solution resistivity inducing an increase of the channel access resistance (Fig. 1 B). The measurement of the access resistance contained several steps that allowed separation of the effects listed above. The planar bilayer membranes were prepared at room temperature by Montal-Mueller technique from solutions of L-Alpha-Diphytanoyl-Lecithin from Avanti Polar Lipids, Inc. (Pelham, AL). Polyethylene glycols (PEGs) of different molecular weights from Aldrich Chem. Co., Inc. (Milwaukee, WI) and dextran 17,900 from Sigma Chemical Co. (St. Louis, MO) were added to 1 M aqueous NaCl solutions (buffered at pH 6.2) to obtain needed weight/weight concentrations. The absence of any significant partition of the polymers, used for the access resistance evaluation was verified by measuring the dependence of the polymerinduced channel conductance change on polymer weight (3). Sufficiently large polymers, namely PEG 3,400; PEG 17,000; and dextran 17,900 were chosen for the access resistance evaluation. The influence of the polymers on dissolved salt (NaCl) activity was examined by a sodium-selective electrode. It was shown that dextran does not change ion activity. In this case, the increase in the total channel resistance is (1)