Hybrid Function of the Na/K-ATPase: Proton Import Accompanying Na/K Exchange

Na/K pumps incessantly extrude 3 Na and retrieve 2 K per ATPase cycle by coupling pump autophosphorylation and autodephosphorylation to conformational changes that alternately expose ion-binding sites to opposite membrane sides. The resulting net outward current monitors cycle turnover rate, limited to a maximum at positive membrane potentials by the slow deocclusion step that releases K to the cytoplasm. At negative potentials, voltage-enhanced rebinding of external Na to Na-selective binding site III slows the cycle by delaying extracellular K access to binding sites I and II. Also at negative potentials, inward proton current flows through Na/K pumps stalled by withdrawal of extracellular K and Na. Though originally viewed as an artifact of such unnatural conditions, this inward proton current persists, albeit diminished, at physiological external [K], [Na], and pH. At physiological nerve and muscle membrane potentials, each Na/K pump is expected to import one proton for every ∼30 completed Na/K transport cycles; at pH 6.0, a proton enters nearly every other cycle. Mutagenesis showed the protons exploit two carboxylates in Na-selective site III and likely a proton wire stabilized by an intervening conserved tyrosine hydroxyl and bound water. We propose proton import involves the rapid, reversible, extracellular Na-releasing conformational transitions that follow the relatively slow deocclusion of the first of the 3 Na ions, which permits site III to become vacant. The inferred co-occurrence of Na/K exchange and proton import during the same conformational cycle identifies Na/K pumps as hybrid transporters, like, e.g., Na-coupled glutamate transporters that also conduct chloride. It remains unclear whether Na/K pump-mediated proton transport has been co-opted for any physiological function. Intriguingly, the predominant AHC(alternating hemiplegia of childhood)-causing mutation of neuronal alpha3 Na/K pumps abolishes site I and II K binding, but leaves proton import unaffected.