Semisynthetic K + channels show that the constricted conformation of the selectivity filter is not the C-type inactivated state

Significance C-type inactivation is a conformational change at the selectivity filter, the ion binding site in a K+ channel that renders it nonconductive. C-type inactivation is important in modulating cellular excitability. Previous studies have suggested a “constricted conformation” for the selectivity filter in the C-type inactivated state. Here, we use protein semisynthesis to introduce unnatural amino acids into the selectivity filter to block it from attaining the constricted conformation. We show that blocking the constricted conformation does not affect C-type inactivation. This study therefore suggests that the constricted conformation of the selectivity filter is not the C-type inactivated state in a K+ channel. The study also highlights ways in which chemical synthesis can be used to manipulate large integral membrane proteins.