title: Only the first and the last hydrophobic segments in the COOH-terminal third of Na,K-ATPase alpha subunit initiate and halt, respectively, membrane translocation of the newly synthesized polypeptide. Implications for the membrane topology.
abstract: We studied the topogenic properties of five hydrophobic segments (H5-H9) in the COOH-terminal third of Na,K-ATPase alpha subunit using in vitro insertion of fusion proteins into endoplasmic reticulum membranes. These fusion proteins consisted of several different lengths of truncated alpha subunit starting at Met729 and a reporter protein, chloramphenicol acetyltransferase, that was linked in frame after each hydrophobic segment. We found that membrane insertion of the newly synthesized COOH-terminal third was initiated by H5 and terminated by H9, indicating that here only H5 and H9 have topogenic function. The other three, H6-H8, did not have topogenic function in the native context and were translocated into the endoplasmic reticulum lumen. These results were in striking contrast to the previous models in which four or six hydrophobic segments were proposed to cross the membrane. Furthermore, the findings suggest a novel mechanism for achieving the final membrane topology of the COOH-terminal third of the alpha subunit.
authors.affiliation: Department of Cell Biology, New York University School of Medicine, New York 10016, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Xie
authors.forename: S A
authors.initials: SA
authors.lastname: Langhans-Rajasekaran
authors.forename: D
authors.initials: D
authors.lastname: Bellovino
authors.forename: T
authors.initials: T
authors.lastname: Morimoto
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D018076:DNA, Complementary; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015398:Signal Transduction; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D010455:Peptides; D011994:Recombinant Proteins; D000254:Sodium-Potassium-Exchanging ATPase