![Figure 2. Electrophiles activate hTRPA1 faster than they react with cytosolic sulfhydryl groups. (A and B) 300 µM H2O2 causes an oxidation-induced increase in roGFP 405/470 ratio in HEK293 (n = 327), prevented by pretreatment with 1 mM NEM (420 s, n = 106) but not by 30 µM IA (60 s, n = 189). Bars, 20 µm. (C) Mean ± SEM H2O2-induced change in 405/470 ratio is reduced by electrophilic stimuli >30 µM IA (60 s; n ≥ 84). *, P < 0.05. (D) IA activates hTRPA1 at [IA] ≥ 1 µM within 120 s. The dashed gray line denotes the response of nontransfected HEK293 to 100 µM IA (first and second stimuli); other colored lines denote hTRPA1-expressing HEK293 cells responding to varying [IA] (first stimulus) and 100 µM IA (second stimulus; n ≥ 36). Colored bars denote the specific duration of each treatment: 1–3 µM (540 s), 10 µM (180 s), and ≥30 µM (120 s). (E) Increasing IA exposure activates hTRPA1 faster, yielding consistent second order reaction rates. (F) Comparison of second order rates for electrophilic modification of GSH (n = 3 each) and TNB (n ≥ 3) and the activation of hTRPA1 (n > 28). (E and F) Error bars denote SEM.](https://cdn.rupress.org/rup/content_public/journal/jgp/147/6/10.1085_jgp.201611581/5/jgp_201611581_fig2.jpeg?Expires=1773520994&Signature=MvEJ1m5eii2zMuAXbMMStQXpJxuWK7bvAJnsjnOhBiAdpheCa4q8863elNdlQ4FTv9QZV98GUrw9iNDShaABmnQhJfJzZtt-1tbScZzAhUaN6Ih0pKqQE45leVHj96qbRUHUlMC-Rf2Ki0Xiar2HzDb1HgdW24KQPqY0NaJrlMmDITcTgLMatzg2xEG00jWRVXk1M09RojOer6o5N4sGQZzvtkiHuUjS7NmRslde1Egkn7bFp6jPMoOykYIIrDdvSQ63RUqFyV6ergyJkpMX1~WEH2LIykwlTBgygbLbQkSrM-OMJFmvSPMTGoy4xFOMTZWNt0vvT~u07AwSUWlWVQ__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Electrophiles activate hTRPA1 faster than they react with cytosolic sulfhydryl groups. (A and B) 300 µM H2O2 causes an oxidation-induced increase in roGFP 405/470 ratio in HEK293 (n = 327), prevented by pretreatment with 1 mM NEM (420 s, n = 106) but not by 30 µM IA (60 s, n = 189). Bars, 20 µm. (C) Mean ± SEM H2O2-induced change in 405/470 ratio is reduced by electrophilic stimuli >30 µM IA (60 s; n ≥ 84). *, P < 0.05. (D) IA activates hTRPA1 at [IA] ≥ 1 µM within 120 s. The dashed gray line denotes the response of nontransfected HEK293 to 100 µM IA (first and second stimuli); other colored lines denote hTRPA1-expressing HEK293 cells responding to varying [IA] (first stimulus) and 100 µM IA (second stimulus; n ≥ 36). Colored bars denote the specific duration of each treatment: 1–3 µM (540 s), 10 µM (180 s), and ≥30 µM (120 s). (E) Increasing IA exposure activates hTRPA1 faster, yielding consistent second order reaction rates. (F) Comparison of second order rates for electrophilic modification of GSH (n = 3 each) and TNB (n ≥ 3) and the activation of hTRPA1 (n > 28). (E and F) Error bars denote SEM.
