Immunoreactivity of subunits of the (Na+ + K+)-ATPase. Cross-reactivity of the alpha, alpha + and beta forms in different organs and species.

The immunologic cross-reactivity of the alpha and alpha + forms of the large subunit and the beta subunit of the (Na+ + K+)-ATPase from brain and kidney preparations was examined using rabbit antiserum prepared against the purified holo lamb kidney enzyme. As previously reported by Sweadner ((1979) J. Biol. Chem. 254, 6060-6067) phosphorylation of the large subunit of the (Na+ + K+)-ATPase in the presence of Na+, Mg2+, and [gamma-32P]ATP revealed that dog and, very likely, rat brain contain two forms of the large subunit (designated alpha and alpha +) while dog, rat, and lamb kidney contain only one form (alpha). The cross-reactivity of the alpha and alpha + forms in these preparations was investigated by resolving the subunits by SDS-polyacrylamide gel electrophoresis. The separated polypeptides were transferred to unmodified nitrocellulose paper and reacted with rabbit anti-lamb kidney serum, followed by detection of the antigen-antibody complex with 125I-labeled protein A and autoradiography. By this method, the alpha and alpha + forms of rat and dog brain, as well as the alpha form found in kidney, were shown to cross-react. In addition, membranes from human cerebral cortex were shown to contain two immunoreactive bands corresponding to the alpha and alpha + forms of dog brain. In contrast, the brain of the insect Manduca sexta contains only one immunoreactive polypeptide with a molecular weight intermediate to the alpha and alpha + forms of dog brain. The beta subunit from lamb, dog and rat kidney and from dog and rat brain cross-reacts with anti-lamb kidney (Na+ + K+)-ATPase serum. The mobility of the beta subunit from dog and rat brain on SDS-polyacrylamide electrophoresis gels is greater than the mobility of the beta subunit from lamb, rat or dog kidney.