Role of the Na,K-ATPaseβ-subunit in the cellular accumulation and maturation of the enzyme as assessed by glycosylation inhibitors

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Abstract
No functional role could yet be established for the glycosylated β-subunit of the Na,K-ATPase. In this study, we describe the intracellular processing of the β-subunit as a glycoprotein in toad bladder cells and the consequences of its structural perturbation with glycosylation inhibitors on the cellular expression of the α- and β-subunits and on the structural and functional maturation of the enzyme. Controlled trypsinolysis of homogenates from pulse-labeled cells reveals that the β-subunit is subjected to glycosylation-dependent structural rearrangements during its intracellular routing. Inhibition of correct terminal glycosylation of the β-subunit with deoxynojirimycin or swainsonine has no effect on the trypsin sensitivity of the α-subunit, its ability to perform cation-dependent conformation changes or the cellular Na,K-ATPase activity. Acquisition of core-sugars is sufficient for the enzyme to assume its catalytic functions. On the other hand, complete inhibition of glycosylation with tunicamycin leads to a destabilization of both the β- and the α-subunits as judged by their higher trypsin sensitivity. In addition, tunicamycin treatment results in a decrease of the amount of newly synthesized β- and α-subunit indicating that a glycoprotein, possibly the β-subunit itself, plays a role in the efficient accumulation of the α-subunit in the endoplasmic reticulum.