To investigate whether nongastric H,K-ATPases transport Na⁺ in exchange for K⁺, and whether different ß isoforms influence their transport properties, we compared the functional properties of human ATP1al1 (AL1), and of the Na,K-ATPase 1 subunit (1) expressed in Xenopus oocytes with different ß subunits. Our results show thatß HK and ß1 NK can produce functional AL1/ß complexes at the oocyte cell surface which, in contrast to 1/ß1 NK and 1/ß HK complexes, exhibit a similar apparent K⁺-affinity. Similar to Na,K-ATPase, AL1/ß complexes are able to decrease intracellular Na⁺ concentrations in Na⁺-loaded oocytes and their K⁺-transport depends on intra- and extracellular Na+ concentrations. Finally, controlled trypsinolysis reveals that ß isoforms influence the protease sensitivity of AL1 and 1 and that AL1/ß complexes, similar to the Na,K-ATPase, can undergo distinct K⁺-, Na⁺- and ouabain-dependent conformational changes. These results provide new evidence that the human nongastric H,K-ATPase interacts with and transports Na⁺ in exchange for K⁺ and that ß isoforms have a distinct effect on the overall structural integrity of AL1, but influence its transport properties less than those of the Na,K-ATPase subunit.