| Abstract
| - Correolide is a novel immunosuppressantthat inhibits the voltage-gated potassium channelKv1.3 [Felix et al. (1999) Biochemistry38, 4922−4930]. [3H]Dihydrocorreolide(diTC) binds with highaffinity to membranes expressing homotetrameric Kv1.3 channels,and high affinity diTC binding can beconferred to the diTC-insensitive channel, Kv3.2, aftersubstitution of three nonconserved residues in S5and S6 with the corresponding amino acids present in Kv1.3 [Hanner et al. (1999) J. Biol. Chem. 274,25237−25244]. Site-directed mutagenesis along S5 and S6 of Kv1.3 was employed to identify thoseresiduesthat contribute to high affinity binding of diTC. Binding of monoiodotyrosine-HgTX1A19Y/Y37F([125I]HgTX1A19Y/Y37F) in the external vestibuleof the channel was used to characterize each mutantfor both tetrameric channel formation and levels of channel expression.Substitutions at Leu346 and Leu353in S5, and Ala413, Val417, Ala421, Pro423, and Val424 in S6, cause the most dramatic effect on diTC bindingto Kv1.3. Some of the critical residues in S6 appear to be present in a region of the protein that alters itsconformation during channel gating. Molecular modeling of the S5−S6 region of Kv1.3 using theX-raycoordinates of the KcsA channel, and other experimental constraints,yield a template that can be used todock diTC in the channel. DiTC appears to bind in the water-filledcavity below the selectivity filter toa hydrophobic pocket contributed by the side chains of specific residues.High affinity binding is predictedto be determined by the complementary shape between the bowl-shapeof the cavity and the shape of theligand. The conformational change that occurs in this region of theprotein during channel gating mayexplain the state-dependent interaction of diTC with Kv1.3.
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