Through the use of osmotic stress, the regulatory effect of chloride ions on hemoglobin function has been reexamined explicitly including water effects. Instead of the 1.6 chloride ions generally assumed linked to oxygen affinity, it now appears that the direct binding of only one ion to the deoxy state is linked, while the rest of the effect is due to the binding of 65 extra water molecules to the oxy state. This result taking into account hydration effects suggests that ligand regulation of """"""""allosteric proteins"""""""" must be re-evaluated with new regard for the activity of water. The peptide alamethicin inserted into bilayer membranes composed of lipids known to form inverted phases of different spontaneous radii showed a clear correlation between the lipid spontaneous curvature and the relative probabilities of different conductance states and therefore channel structures. The dependence of ionic channel expression on the packing strain in lipid bilayer revealed by these experiments suggests an active role for membrane lipid composition in regulating membrane protein activity. Currents through fully open single channels formed by Staphylococcus aureus alpha toxin were subjected to fluctuation analysis. A new mechanism of excess noise generation in an ion channel was identified and related to a reversible ionization of residues in the channel-forming molecule. The reaction parameters and the number of residues participating in the ionization process were extracted. The ability of noise analysis to study reactions within a single ionic channel demonstrates the potential power of the technique as structural tool for channel function analysis.
