期刊
PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY
卷 461, 期 6, 页码 665-675出版社
SPRINGER
DOI: 10.1007/s00424-011-0924-7
关键词
Potassium channel; Patch clamp; Ion channel; Electrophysiology; Calcium-activated potassium channel
类别
资金
- The Medical Research Council
- The British Heart Foundation
The large conductance, voltage- and calcium-activated potassium channel, BKCa, is a known target for the gasotransmitter, carbon monoxide (CO). Activation of BKCa by CO modulates cellular excitability and contributes to the physiology of a diverse array of processes, including vascular tone and oxygen-sensing. Currently, there is no consensus regarding the molecular mechanisms underpinning reception of CO by the BKCa. Here, employing voltage-clamped, inside-out patches from HEK293 cells expressing single, double and triple cysteine mutations in the BKCa alpha-subunit, we test the hypothesis that CO regulation is conferred upon the channel by interactions with cysteine residues within the RCK2 domain. In physiological [Ca2+](i), all mutants carrying a cysteine substitution at position 911 (C911G) demonstrated significantly reduced CO sensitivity; the C911G mutant did not express altered Ca2+-sensitivity. In contrast, histidine residues in RCK1 domain, previously shown to ablate CO activation in low [Ca2+](i), actually increased CO sensitivity when [Ca2+](i) was in the physiological range. Importantly, cyanide, employed here as a substituent for CO at potential metal centres, occluded activation by CO; this effect was freely reversible. Taken together, these data suggest that a specific cysteine residue in the C-terminal domain, which is close to the Ca2+ bowl but which is not involved in Ca2+ activation, confers significant CO sensitivity to BKCa channels. The rapid reversibility of CO and cyanide binding, coupled to information garnered from other CO-binding proteins, suggests that C911 may be involved in formation of a transition metal cluster which can bind and, thereafter, activate BKCa.
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