Reconstitution of ion channels.

Abstract

The first model of the structure of cell membranes was developed very early from indirect evidence.' It postulated the existence of a bimolecular lipid membrane which serves as the main and electrical isolating part of the cell membrane. The proteins were either associated with or incorporated into the membrane.Z Following this idea, it was postulated that the proteins being incorporated into the bimolecular lipid matrix (integral membrane proteins) were the source of ion transport across the electrical isolating lipid membrane.3 Ion channels formed by these proteins were one of the mechanisms being considered to be involved in the ion transport across cell membranes. These ion channel-forming proteins were proposed to be involved in the electrical excitation of biological membranes very early.4 For a long time, however, it was impossible to investigate directly the properties of these channels, i.e., in single channel experiments. Limited by the low current and time resolution of electrophysiological measurements, only multichannel experiments were possible. Due to the fact that multichannel data can be explained by a larger number of single channel models there was a need for detailed single channel data to obtain a more in-depth view of channel function. In addition, all of the single channel parameters calculated from the electrophysiological experiments were scattered over a wide range of values. Nevertheless, from electrophysiological experiments a large number of different ion channels were postulated to be active in biological membranes, and highly specific properties were given to these hypothetical ion channels. However, until 1962 not even the existence of single ion channels could be shown experimentally. Following that line of thought, the first technique for the formation of planar lipid bilayers was established in 1962 by Mueller et al.,' using large amounts of organic solvents to stabilize the artificial membrane. The first single channel fluctuations shown were those of EIM and gramicidin in planar lipid bilayers which were formed according to that t echn iq~e .~ -~ For a period of time, single channel fluctuations being induced by different polypeptide antibiotics and related substameslo in planar bilayers were the only really measured single channel fluctuations. At that time it was not possible to measure current fluctuations of single ion channels in cell membranes directly. It was obvious that the incorporation of channel-forming proteins into planar lipid bilayers would be a useful technique for finding the proof of the existence of single ion channels in biological membranes and for investigating the properties of these hypothetical channels in singleand multichannel experiments. Most of the early reconstitution experiments were not very successful. Problems occurred because these bilayers contained large amounts of organic solvents, techniques had to be developed to incorporate C ri tic al R ev ie w s in B io ch em is tr y an d M ol ec ul ar B io lo gy D ow nl oa de d fr om in fo rm ah ea lth ca re .c om b y 41 .2 20 .2 8. 51 o n 05 /2 0/ 14