This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The extracellular entryway of the bacterial potassium channel of Streptomyces lividans (KcsA) is homologous to eukaryotic voltage gated channels. For this reason, KcsA is used as a template for the binding of extracellular pore blockers. Animal venoms such as snake and scorpion venoms are de facto libraries of naturally occurring toxins. The venom from the Snake Dendroaspis angusticeps or from the Scorpion Leiurus quinquestriatus hebraeus was screened against immobilized KcsA using affinity chromatography. Following extensive washes, the channel was eluted along with specifically bound toxins. MALDI-TOF MS was used as a tool to quickly identify small protein toxins from their molecular mass. This approach provides a rapid method for identifying potential inhibitors of eukaryotic potassium channels. We are developing methods for rapidly determining the primary sequences of newly discovered channel-binding toxins.
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