title: Human cardiac potassium channel DNA polymorphism modulates access to drug-binding site and causes drug resistance.
abstract: Expression of voltage-gated K channel, shaker-related subfamily, member 5 (KCNA5) underlies the human atrial ultra-rapid delayed rectifier K current (I(Kur)). The KCNA5 polymorphism resulting in P532L in the C terminus generates I(Kur) that is indistinguishable from wild type at baseline but strikingly resistant to drug block. In the present study, truncating the C terminus of KCNA5 generated a channel with wild-type drug sensitivity, which indicated that P532 is not a drug-binding site. Secondary structure prediction algorithms identified a probable alpha-helix in P532L that is absent in wild-type channels. We therefore assessed drug sensitivity of I(Kur) generated in vitro in CHO and HEK cells by channels predicted to exhibit or lack this C-terminal alpha-helix. All constructs displayed near-identical I(Kur) in the absence of drug challenge. However, those predicted to lack the C-terminal alpha-helix generated quinidine-sensitive currents (43-51% block by 10 microM quinidine), while the currents generated by those constructs predicted to generate a C-terminal alpha-helix were inhibited less than 12%. Circular dichroism spectroscopy revealed an alpha-helical signature with peptides derived from drug-resistant channels and no organized structure in those associated with wild-type drug sensitivity. In conclusion, we found that this secondary structure in the KCNA5 C terminus, absent in wild-type channels but generated by a naturally occurring DNA polymorphism, does not alter baseline currents but renders the channel drug resistant. Our data support a model in which this structure impairs access of the drug to a pore-binding site.
authors: Division of Clinical Pharmacology, Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
authors: Benoit
authors: B
authors: Drolet
authors: Chantale
authors: C
authors: Simard
authors: Laura
authors: L
authors: Mizoue
authors: Dan M
authors: DM
authors: Roden
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics*; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004351:Drug Resistance / Q000235:genetics*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D051666:Kv1.5 Potassium Channel; D008564:Membrane Potentials / Q000187:drug effects; D011110:Polymorphism, Genetic; D024642:Potassium Channels, Voltage-Gated / Q000235:genetics* / Q000378:metabolism; D017433:Protein Structure, Secondary / Q000235:genetics; D011802:Quinidine / Q000494:pharmacology*
chemical_list: D004791:Enzyme Inhibitors; C496981:KCNA5 protein, human; D051666:Kv1.5 Potassium Channel; D024642:Potassium Channels, Voltage-Gated; D011802:Quinidine