Shaker potassium channels undergo a slow C-type inactivation which can be hastened dramatically by single-point mutations in or near the pore region. We found that the oxidizing agent chloramine-T (Chl-T) causes an irreversible loss of current for those mutants which show C-type inactivation. For several mutants at position T449, which show a wide spectrum of inactivation time constants, the time constant of current rundown induced by Chl-T correlated with the speed of inactivation. Rundown was accelerated when the channels were in the inactivated state but rundown also occurred when channels were not opened or inactivated. Apparently, only those channels which can undergo C-type inactivation are accessible to Chl-T. In order to gain information about the target amino-acid residue for the action of Chl-T and the structural rearrangements occurring during C-type inactivation, several mutant channel proteins were compared with respect to their response to Chl-T. Since Chl-T can oxidize cysteine and methionine residues, we mutated the possible targets in and close to the pore region, namely C462 to A, and M440 and M448 to I. While the residues M440 and C462 were not important for channel rundown, mutation of M448 to I made the channels more resistant to Chl-T by about one order of magnitude. While inactivation was accelerated upon application of Chl-T in most mutants, mutation of M448 to I abolished this effect on the time course of inactivation, indicating that M448 is one of the target residues for Chl-T.