Abstract
Tryptic cleavage of the catalytic subunit of kidney Na,K-ATPase in the E1 conformation effects a change in kinetic behavior apparent at low ATP concentration. Thus, at < or = 10 microM ATP, K+ inhibits Na(+)-dependent ATPase activity of the undigested enzyme but activates activity of the digested enzyme. With time of trypsinolysis, a transient increase followed by a decrease in activity is observed at low [ATP], whereas at high [ATP] (1 mM), activity is progressively reduced. At low [ATP], the trypsin-treated/control activity ratio was > or = 3-fold higher with K+ compared to the ratio observed with the K+ congener Li+. Also, the relative Na/K exchange activity (22Na+ influx into K(+)-loaded inside-out vesicles from erythrocytes) with either 0.01 mM ATP or 1 mM CTP compared to 1 mM ATP was greater for the trypsin-treated than for the control enzyme. The kinetic change is correlated with the initial rapid cleavage of the N-terminal tryptic fragment (< or = 30 residues) from the catalytic subunit. It is concluded that this segment regulates the K+ deocclusion pathway of the reaction; removal of this fragment produces a modified active species having an increased rate of K+ deocclusion.