ABSTRACT
The currently accepted topographical model for the organization of the alpha-subunit of the Na+, K+-ATPase in the membrane considers that the protein has ten transmembrane segments and six cytoplasmic loops. Evidence of interaction between the cytoplasmic regions may contribute to a better understanding of the structure/function relationship of this protein. In this study, the first four cytoplasmic segments (C1, C2, C3 and C4) of the rat alpha1 subunit were expressed in Escherichia Coli. The large cytoplasmic loop between transmembrane segments four and five (C3) retained its native structure as demonstrated by the ability of ATP to protect against chemical modification by Fluorescein 5-isothiocyanate (FITC). Interaction studies were conducted by an overlay assay (Far Western blots) and surface plasmon resonance technology. We observed that C3 interacts with the N-terminal segment of the Na+, K+-ATPase, C1; and that both C1 and C3 interact with the cytoplasmic segments C2 and C4.
