Preferential closed channel blockade of HERG potassium currents by chemically synthesised BeKm-1 scorpion toxin

• Published in: FEBS letters 2003-07, Vol.547 (1), p.20-26
• Main Authors: Milnes, James T, Dempsey, Christopher E, Ridley, John M, Crociani, Olivia, Arcangeli, Annarosa, Hancox, Jules C, Witchel, Harry J
• Format: Article
• Language: English
• Subjects: ANOVA, analysis of variance; BeKm-1; Buthus eupeus; Cation Transport Proteins; Cell Line; Centruroides noxius; CI, 95% confidence intervals; DNA-Binding Proteins; DQF-COSY, double quantum filtered correlation spectroscopy; ERG1 Potassium Channel; Ergtoxin; Ether-A-Go-Go Potassium Channels; Fmoc, fluorenylmethoxycarbonyl; h, Hill coefficient; HEK, human embryonic kidney; HERG, human ether-a-go-go-related gene; HPLC, high performance liquid chromatography; Human ether-a-go-go-related gene; Humans; I HERG, current mediated by the HERG channel; I Kr; IC50, half-maximal inhibitory drug concentration; IKr; k, slope factor describing voltage-dependent activation; Kinetics; Patch-Clamp Techniques; Potassium Channel Blockers - chemical synthesis; Potassium Channel Blockers - pharmacology; Potassium Channels - drug effects; Potassium Channels - genetics; Potassium Channels - physiology; Potassium Channels, Voltage-Gated; Protein Folding; Rapid delayed rectifier; rBeKm-1, recombinant BeKm-1; Recombinant Proteins - antagonists & inhibitors; S-to-S, start-to-start interval; Scorpion toxin; Scorpion Venoms - chemical synthesis; Scorpion Venoms - chemistry; Scorpion Venoms - pharmacology; Thermodynamics; Trans-Activators; Transcriptional Regulator ERG; Transfection; V 0.5, half-maximal activation voltage
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/S0014-5793(03)00662-8

The scorpion toxin peptide BeKm-1 was synthesised by fluorenylmethoxycarbonyl solid phase chemistry and folded by air oxidation. The peptide’s effects on heterologous human ether-a-go-go-related gene potassium current ( I HERG) in HEK293 cells were assessed using ‘whole-cell’ patch clamp. Blockade of I HERG by BeKm-1 was concentration-dependent, temperature-dependent, and rapid in onset and reversibility. Blockade also exhibited inverse voltage dependence, inverse dependence on duration of depolarisation, and reverse use- and frequency-dependence. Blockade by BeKm-1 and recombinant ergtoxin, another scorpion toxin known to block HERG, differed in their recovery from HERG current inactivation elicited by strong depolarisation and in their ability to block HERG when the channels were already activated. We conclude that synthetic BeKm-1 toxin blocks HERG preferentially through a closed (resting) state channel blockade mechanism, although some open channel blockade also occurs.