Inhibition of Large-Conductance Ca2+-Activated K+ Channels by Nanomolar Concentrations of Ag+

Abstract

Silver has been widely used in various medical products because of its antibacterial properties. However, there is only limited information concerning silver-related cytotoxicity. In this study we show that Ag ϩ at low nanomolar concentrations (Ͻ10 nM) strongly inhibits the activity of large-conductance Ca 2ϩ -activated K ϩ channels (BK) (Slo1), a widely expressed and physiologically important potassium channel. The Ag ϩ inhibition is caused by irreversible modification on cytosolically accessible parts of the BK channel. At least four intracellular cysteines are involved in this process. In addition, at least one of these key cysteines is not accessible to the bulkier thiolateactive reagent [2-(trimethylammonium)ethyl] methanethiosulfonate bromide. One of the cysteine-less constructs generated in this study shows gating properties similar to wild-type BK channel but with much lower Ag ϩ sensitivity, in which the Ag ϩ modification rate was decreased by approximately 20-fold. The results from the present study suggest a possible contribution of BK channel inhibition to the cytotoxicity of Ag ϩ in humans and other species.

Charge substitution for a deep-pore residue reveals structural dynamics during BK channel gating

“…For nAChR, the linear free energy analysis has yielded a tremendous amount of insight into the structural details of the arrangement of the pore and how nAChR channels gate ( Grosman et al, 2000 ; Cymes et al, 2002 ), as did the proton-probing method ( Cymes et al, 2005 ; Cymes and Grosman, 2008 , 2011 ). For BK channels, because of a large number of endogenous cysteines, the cysteine scanning approach is problematic ( Tang et al, 2001 ; Zhang and Horrigan, 2005 ; Zhou et al, 2010 ). As we found out in this study, BK channels, possibly because of their large pore size ( Li and Aldrich, 2004 , 2006 ; Brelidze and Magleby, 2005 ; Wilkens and Aldrich, 2006 ; Tang et al, 2009 ), are quite tolerant for large side-chain substitutions in the pore.…”

Closed state-coupled C-type inactivation in BK channels

“…The observation that permeant ions, such as thallium, can affect BK channel activation further led to the notion that the BK channel's activation gate may exist near or at the selectivity filter (21). A direct determination of the state dependence of ion accessibility at the selectivity filter using small cysteine modification reagents like cadmium and silver, as previously done in CNG channels (22), has been hampered by the complexity of BK channels, which have numerous endogenous cysteine residues whose modifications affect channel functions (23,24). Thus, it is important to explore alternative approaches to localizing the activation gate in BK channels.…”

Silver ions enhance UVB‐induced phosphorylation of histone H2AX

“…Ag + was found to affect the cell viability and morphology of skin fibroblasts and keratinocytes [Hidalgo et al, ]. Zhou et al showed that Ag + strongly inhibited the properties of Ca 2+ ‐activated K + channels, a widely expressed and physiological important channel [Zhou et al, ]. Furthermore, Ag + induced oxidative stress through the generation of reactive oxygen species and/or by impairing glutathione, an important cellular antioxidant [Cortese‐Krott et al, ].…”

“…Additional Supporting Information may be found in the online version of this article. Grant sponsor: JSPS KAKENHI; Grant numbers: 24510084 and 25340048. strongly inhibited the properties of Ca 21 -activated K 1 channels, a widely expressed and physiological important channel [Zhou et al, 2010]. Furthermore, Ag 1 induced oxidative stress through the generation of reactive oxygen species and/or by impairing glutathione, an important cellular antioxidant [Cortese-Krott et al, 2009].…”