A 19KD C-Terminal Tryptic Fragment of the Alpha Chain of Na/K-ATPase, Essential for Cation Occlusion and Transport

Abstract

Trypsin, in the presence of Rb but absence of Ca, digests off half the protein of pig renal Na/K-ATPase leaving a 19KD membrane-embedded fragment of the alpha chain with almost normal Rb and Na occlusion, but no ATP-dependent functions. Subsequent digestion, now in the absence of Rb but the presence of Ca, leads to rapid loss of the 19KD peptide and parallel loss of cation occlusion. Thus the peptide is essential for occlusion. Its N-terminal sequence is NPKTDKLVNERLISMA. It begins at residue 830 and extends towards the C-terminus. 19KD-containing membranes can reconstitute into phospholipid vesicles, to sustain a slow Rb-Rb exchange. Cation occlusion sites and the transport pathway, within trans-membrane segments, are thus quite separate from the ATP binding site, located on the cytoplasmic domain of the alpha chain.