Received for publication May 17, 2004.
Revised June 22, 2004.
Accepted for publication July 27, 2004.

Mapping of maurotoxin binding sites on hKv1.2, hkv1.3 and hIKCa1 channels

Abstract

Maurotoxin (MTX) is a potent blocker of human voltage-activated Kv1.2 and intermediate-conductance calcium-activated potassium channels, hIKCa1. Since its blocking affinity on both channels is similar, although the pore region of these channels show only few conserved amino acids, we aimed to characterize the binding sites of MTX in these channels. Investigating the pH_o dependency of MTX block on current through hKv1.2 channels, we concluded that the block is less pH_o sensitive than for hIKCa1 channels. Using mutant cycle analysis and computer docking we tried to identify the amino acids through which MTX binds to hKv1.2 and hIKCa1 channels. We report that MTX interacts with hKv1.2 mainly though six strong interactions. Lys²³ from MTX protrudes into the channel pore interacting with the GYGD motif, while Tyr³² and Lys⁷ interact with Val³⁸¹, Asp³⁶³, and Glu³⁵⁵, stabilising the toxin onto the channel pore. Since only Val³⁸¹, Asp³⁶³, and the GYGD motif are conserved in hIKCa1 channels, and the replacement of His³⁹⁹ from hKv1.3 channels with a threonine makes this channel MTX sensitive, we concluded that MTX binds to all three channels through the same amino acids, Glu³⁵⁵, although important, being not essential in MTX recognition. A negatively charged amino acid in this position could better stabilize the toxin-channel interaction, and could explain the pH_o sensitivity of MTX block on current through hIKCa1 versus hKv1.2 channels.

Key words: Structure-activity relationships and modeling, Func. analysis receptor/ion channel mutants, Mutagenesis/Chimeric approaches, Structure/function/mechanism

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