Modification of the cation selectivity filter and the calcium receptor of the Ca-stimulated K channel in resealed ghosts of human red blood cells by low levels of incorporated trypsin.

The transport properties of the Ca-activated, K channel in the resealed human blood cell ghost can be modified by the action of incorporated trypsin. Membranes were maximally depleted of diffusible cytoplasmic components by hemolysis on a gel filtration column at 0 degree C. Subsequently, isotonicity was restored and 0.01-1 microgram/ml trypsin incorporated. Partial digestion of the membrane proteins occurred during resealing. As the degree of tryptic digestion increased, the channel became initially permeable to K and later to both and Na; and then the channel became refractory to the action of applied Ca. The observations suggest that tryptic digestion of proteins at the inner membrane surface leads to modifications of the selectivity filter and the Ca-receptor site of the channel. The modifications probably stem from alterations at the inner surface of a transmembrane protein which acts as a channel. Under conditions where selectivity is lost, the channel is still inhibited by externally applied TbCl3 .