State dependence of disulfide bond formation in mutants R198C and R201C expressed in Xenopus oocytes. (A) Representative current traces of mutant R198C before or after Cu-Phen application in the channel closed state and after washout with 1 mM DTT. Oocytes were incubated for 2 min at −80 mV in ND96 solution containing 100 µM Cu-Phen, washed with ND96 for another 5 min at −80 mV, and then depolarized as in Fig. 4 A; subsequently, the oocytes were washed out in the presence of 1 mM DTT and subjected to the same depolarizing protocol; n = 7. (B) Representative current traces of mutant R198C before or after Cu-Phen application in the channel open state. From a holding potential of −80 mV, oocytes were subjected to a depolarizing step to 0 mV. 2 s after the beginning of the test pulse, fast application of 100 µM Cu-Phen was applied for up to 2 min; n = 7. (C) Representative current traces of mutant R201C before or after Cu-Phen application at −80 mV and after washout with 1 mM DTT. Oocytes were treated and currents were evoked as in Fig. 4 A; n = 7. (D) Representative current traces of mutant R201C before or after Cu-Phen application in the channel open state. Oocytes were treated and currents were evoked as in B; n = 10.