Functional identification of ion binding sites at the internal end of the pore in Shaker K+ channels

The inner end of the pore in voltage-gated K+ channels is the site of conformational changes related to gating and contains binding sites for permeant ions and pore-blocking molecules including quaternary ammonium ions and drugs. In order to determine the location and affinity of ion binding sites we probed the Shaker K+ channel with the quaternary ammonium analogue, tetrabutyl antimony (TBSb), a compound that is sufficiently electron dense to have been observed to occupy the cavity site in the bacterial K+ channel, KcsA. TBSb has K+ channel blocking properties analogous to those of tetrabutyl ammonium (TBA), and kinetics slow enough to be reliably measured. In the presence of external TEA, the internal TBSb on-rate decreased with increased internal K+ concentration as if these permeant ions prevented TBSb access to its site in the pore. The TBSb off-rate in low K+ was increased with external TEA addition and then reduced with increased internal K+. We found several differences between the behaviour of internal TBSb and TEA suggesting these molecules bind to distinct but interacting sites in the pore. We also found several differences in how K+ and Rb+ ions occupy sites in the inner end of the pore. These data suggest the presence of three sites in the inner end of the pore: (1) a site near the cytoplasmic end that binds TEA and K+ (but not Rb+) ions; K+ ions binding to this site inhibit TBSb exit from the pore; (2) a TBSb site slightly more into the pore that is rarely occupied by K+ or Rb+ ions; (3) a site further into the pore that has a high affinity for K+ and Rb+ ions; occupancy of this site by these permeant ions increases the TBSb off-rate. These results provide information on the fine-structure of ion interactions with the inner end of the pore in K+ channels

A Tool for Inverse Modeling of Spectral Measurements in Deep and Shallow Waters

A software tool was developed for simulating and inverse modeling of optical spectral measurements in deep and shallow waters above and below the water surface. It supports eight major spectrum types which are commonly measured by instruments on ship: irradiance reflectance, remote sensing reflectance, downwelling irradiance, upwelling radiance, absorption, attenuation, specular reflectance at the surface, and bottom reflectance. Calculation is based on analytical models. For deep water different well-established models are included, for shallow water and surface reflections new models were developed. The program is designed as a user-friendly, sensor-independent spectra generator and spectra analyzer with well documented calculation steps and automatic result visualization. It is suited to generate and analyze large series of spectra. All model constants and input spectra can be changed easily for adaptation to a specific region. This contribution summarizes the models, explains the inversion techniques, and describes how to apply the program