The partial reactions of the Na(+)- and Na(+) + K(+)-activated adenosine triphosphatases.

Kinetic investigations carried out in a number of laboratories have accumulated evidence favoring modification of the Albers-Post mechanism. The results of the rapid mixing studies involving the eel enzyme indicate that the complex kinetic behavior is confined to the Na(+)-activated reaction pathway (Na-ATPase). The main conceptual problem in interpreting the dephosphorylation experiments involves the intermediate component, which turns over too slowly to account for the overall velocity of Pi production in the presence of Na+ and K+ and exhibits behavior compatible with an ADP-insensitive phosphoenzyme. Attempts to simulate the dephosphorylation reaction using schemes in which the intermediate component represents a precursor to the K(+)-sensitive phosphoenzyme, E2P, were unsuccessful in reproducing both the pre-steady-state and steady-state time dependence. When Na+ and K+ were both present during phosphorylation, the time course of dephosphorylation showed no evidence of an intermediate decay component, implying that K+ either prevents its formation or accelerates its turnover. Complex kinetic behavior was also observed in the phosphorylation reaction under conditions where the reaction was initiated by the simultaneous addition of ATP, Na+, and Mg2+. Preincubation with Na+ eliminated the biexponential pattern of accumulation so that only the fast phase was seen. The proportion of EP in the slow phase of phosphorylation was approximately equal to the fraction of EP in the intermediate phase of dephosphorylation (roughly one-third of the sites), suggesting that the two may be related to the same catalytic activity. To try to explain these observations using recent modifications to the Albers-Post mechanism is difficult without invoking additional complex effects of the transported ions. We propose that a series model for phosphorylation is inadequate and that further modification of the mechanism is required. The alternative to a consecutive mechanism is a parallel pathway scheme: [sequence: see text] In this model the enzyme exists in two distinct forms which are distributed in the upper and lower pathways in a ratio of 2:1. In the lower pathway the rates of phosphorylation and E2P hydrolysis are controlled by the kinetics of ligand binding because of a structural constraint (ion channel?) imposed by the transport protein. When phosphorylation is carried out in the presence of Na+ alone, E2P and E2P' accumulates rapidly and give rise to the fast and intermediate components of dephosphorylation, respectively. Preincubation with Na+ and K+ eliminates the functional differences between these pathways by removing the kinetic dependence of ligand binding, resulting in behavior that conforms to the Albers-Post mechanism.(ABSTRACT TRUNCATED AT 400 WORDS)