A Model for the Homotypic Interaction between Na(+),K(+)-ATPase β(1) Subunits Reveals the Role of Extracellular Residues 221–229 in Its Ig-Like Domain

The Na(+), K(+)-ATPase transports Na(+) and K(+) across the membrane of all animal cells. In addition to its ion transporting function, the Na(+), K(+)-ATPase acts as a homotypic epithelial cell adhesion molecule via its β(1) subunit. The extracellular region of the Na(+), K(+)-ATPase β(1) subunit includes a single globular immunoglobulin-like domain. We performed Molecular Dynamics simulations of the ectodomain of the β(1) subunit and a refined protein-protein docking prediction. Our results show that the β(1) subunit Ig-like domain maintains an independent structure and dimerizes in an antiparallel fashion. Analysis of the putative interface identified segment Lys221-Tyr229. We generated triple mutations on YFP-β(1) subunit fusion proteins to assess the contribution of these residues. CHO fibroblasts transfected with mutant β(1) subunits showed a significantly decreased cell-cell adhesion. Association of β(1) subunits in vitro was also reduced, as determined by pull-down assays. Altogether, we conclude that two Na(+), K(+)-ATPase molecules recognize each other by a large interface spanning residues 221–229 and 198–207 on their β(1) subunits.