A point mutation in the maxi-K clone dSlo forms a high affinity site for charybdotoxin

作者：Myers MP^*; Stampe P

来源：Neuropharmacology, 2000, 39(1): 11-20.

DOI：10.1016/S0028-3908(99)00074-X

• potassium channel
• Ca2+-activated K+ channel
• CTX
• Slo
• hSlo
• mSlo

This work investigated the interaction of CTX with two cloned analogues of the maxi-K channel, dSlo and hSlo. dSlo has been reported to be CTX insensitive. Single channel analysis revealed that dSlo was weakly blocked by the toxin, with a very high K-D of 5.8 mu M. The hSlo channels bound wild-type, recombinant CTX with high affinity and in a bimolecular fashion, and displayed a half-blocking concentration (K-D) of 36 nM. A glutamate residue was substituted for the wild-type threonine at position 290 in dSlo. The mutant channel was expressed in COS-7 cells and reconstituted into lipid bilayers for single channel analysis. The mutant channel bound wild-type, recombinant CTX with high affinity and in a bimolecular fashion, and displayed a half-blocking concentration (K-D) of 23 nM. Changing just one residue from threonine to glutamate at position 290 in dSlo changed the affinity of the channel's CTX-receptor over 100-fold. Kinetic analysis revealed that this large increase in affinity was due to decreasing the dissociation rate of the toxin. These results suggest that a CTX receptor does exist in the dSlo channel mouth and that the threonine at position 290 destabilizes the toxin on the binding site.

• 出版日期2000