Grenerating a high affinity scorpion toxin receptor in KcsA-Kv1.3 chimericpotassium channels

The crystal structure of the bacterial K+ channel, KcsA (Doyle, D. A., Mora is, C. J., Pfuetzner, R. A., Kuo, A., Gulbis, J. M., Cohen, S. L., Chait, B . T., and MacKinnon, R. (1998) Science 280, 69-77), and subsequent mutagene sis have revealed a high structural conservation from bacteria to human (Ma cKinnon, R., Cohen, S, L., Kuo, A., Lee, A., and Chait, B. T. (1998) Scienc e 280, 106-109). me have explored this conservation by swapping subregions of the M1-M2 linker of KcsA with those of the S5-S6 linker of the human Kv- channel Kv1.3. The chimeric K+ channel constructs were expressed in Escheri chia coil, and their multimeric state was analyzed after purification. We u sed two scorpion toxins, kaliotoxin and hongotoxin 1, which bind specifical ly to Kv1.3, to analyze the pharmacological properties of the KcsA-Kv1.3 ch imeras. The results demonstrate that the high affinity scorpion toxin recep tor of Kv1.3 could be transferred to KcsA. Our biochemical studies with pur ified KcsA-Kv1.3 chimeras provide direct chemical evidence that a tetrameri c channel structure is necessary for forming a functional scorpion toxin re ceptor. We have obtained KcsA-Kv1.3 chimeras with kaliotoxin affinities (IC 50 values of similar to 4 pM) like native Kv1.3 channels. Furthermore, we s how that a subregion of the S5-S6 linker may be an important determinant of the pharmacological profile of K+ channels. Using available structural inf ormation on KcsA and kaliotoxin, we have developed a structural model for t he complex between KcsA-1.3 chimeras and kaliotoxin to aid future pharmacol ogical studies of K+ channels.