The characteristics of potassium channel block by a diverse group of quaternary ammonium (QA) ions was examined in squid axons. Altering the size and nature of the head and/or tail groups of the QA ions applied internally produced only quantitative differences in the potassium current block. Although their entry rate is diminished, compounds with head groups as large as 11 X 12 A are capable of occluding the channel, whereas the smallest QA ions, with head groups approximately 5 X 6 A, are not potent blockers. When one or three terminal hydrogens of the head group were replaced by hydroxyl moieties, the compound's blocking ability was diminished, suggesting that QA binding is not improved by hydrogen bonding at these positions. QA ions bound to their site within the potassium channel with 1:1 stoichiometry, and the site is perhaps 20% or more of the distance through the membrane electric field. Raising external potassium concentration did not alter the steady-state or kinetic features of the QA block of outward potassium currents; however, increasing temperature or adding Ba2+ internally increased the rate of decay of the QA-blocked currents. From the structure-function analysis of the QA ions, projections concerning both the architecture of the potassium channel's inner mouth and the significance of various chemical constituents of the ions were made. The potassium channel may now be pictured as having a wider mouth (up to 11 X 12 A) extending to the QA binding site and then narrowing quickly to the region of channel selectivity. Important alterations that improve the blocking ability of the compounds include: (a) lengthening the alkyl hydrocarbon tail group (up to 10 carbon), (b) lengthening a second hydrocarbon chain of the head group (e.g., decyldimethylphenylammonium bromide [C10DM phi]), and (c) adding a carbonyl moiety to the tail (e.g., ambutonium).