In the 1970s, it became apparent that biological membranes of cells incorporate nanoscopic channels composed of proteins that are embedded in lipid bilayers. The function of the channels is to provide a gated pore that allows solutes such as sodium, potassium, and calcium ions to cross the otherwise impermeable lipid bilayer barrier that surrounds all cells. Artificial bilayers are readily produced by self-assembly of amphiphilic compounds such as phospholipids. For instance, bilayers of phospholipid 5 nm thick can be produced as membranes covering a 100 micrometer diameter hole in a thin plastic sheet. Such membranes have dimensions appropriate for nanopore analysis, provided that a pore can be produced in the bilayer.
Hladky and Haydon  showed that the bacterial antibiotic gramicidin spontaneously inserted into lipid bilayers and could form continuous channels that conducted ionic current. Gramicidin, with a pore diameter of ~0.4 nm, is the smallest useful nanopore. The gramicin pore contains a chain of approximately 10 water molecules that form a single hydrogen-bonded strand across the bilayer. In the presence of a voltage, the pore conducts an ionic current of a few picoamperes, the amplitude depending on the voltage and ion concentration. The gramicidin pore is too small to be used directly to detect solutes other than ions, but Cornell and co-workers  have shown that it can be used as a biosensor if it is coupled to a specific antibody. When a target antigen binds to the antibody, the ability of the gramicidin pore to conduct ionic current is inhibited. The reduction in current can then be used to estimate the presence and concentration of the antigen.
The fact that such pores could conduct polar and ionic solutes across a membrane suggested that a larger pore might be used to detect macromolecules in solution. This concept was first tested by Bezrukov et al.  who showed that ionic current through the alpha hemolysin pore was affected by the presence of polyethylene glycol (PEG). The current fluctuations appeared only at certain molecular size ranges: PEG molecules that were smaller or larger than the pore had little effect, while PEG in the size range of the pore (0.5-1.5 nm) produced large current fluctuations. This allowed the authors to conclude that PEG could enter the pore and affect ionic current. Because PEG has no net electrical charge, the molecules enter the pore by diffusion, rather than by an energy-dependent process. (See Bezrukov  for review.)
At about the same time, Walker and co-workers  demonstrated that the hemolysin pore could be modified to serve as a metal biosensor. In this work, the hemolysin was modified by introducing histidine for amino acids 130-134 in the central glycine-rich loop of the protein. The modified pore had a conductance that could be modulated by micromolar concentrations of ions such as zinc, suggesting that modified protein pores such as hemolysin could serve as sensitive metal ion sensors.
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