title: Stop-and-move of a marginally hydrophobic segment translocating across the endoplasmic reticulum membrane.
abstract: Many membrane proteins are cotranslationally integrated into the endoplasmic reticulum membrane via the protein-conducting channel, the so-called translocon. The hydrophobic transmembrane segment of the translocating nascent polypeptide chain stops at the translocon and then moves laterally into the membrane. Partitioning of the hydrophobic segment into the membrane is the primary determinant for membrane insertion. Here, we examined the behavior of a marginally hydrophobic segment at the translocon and found that its stop-translocation was greatly affected by the C-terminally attached ribosomes. The marginally hydrophobic segment first stops at the membrane and then moves into the lumen as long as the nascent chain is attached to translating ribosomes. When it is released from the ribosome by the termination codon, the marginally hydrophobic segment does not move. Puromycin or RNase treatment also suppressed movement. The movement was reversibly inhibited by high-salt conditions and irreversibly inhibited by ethylenediaminetetraacetic acid. There is an unstable state prior to the stable membrane insertion of the transmembrane segment. This characteristic state is maintained by the synthesizing ribosome.
authors.affiliation: Graduate School of Life Science, University of Hyogo, Kouto Ako-gun, Hyogo 678-1297 Japan.
authors.forename: Yukiko
authors.initials: Y
authors.lastname: Onishi
authors.forename: Marifu
authors.initials: M
authors.lastname: Yamagishi
authors.forename: Kenta
authors.initials: K
authors.lastname: Imai
authors.forename: Hidenobu
authors.initials: H
authors.lastname: Fujita
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Kida
authors.forename: Masao
authors.initials: M
authors.lastname: Sakaguchi
mesh_terms: D000818:Animals; D004721:Endoplasmic Reticulum / Q000235:genetics / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D007425:Intracellular Membranes / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport; D011817:Rabbits; D012270:Ribosomes / Q000235:genetics / Q000378:metabolism
chemical_list: D008565:Membrane Proteins
keywords: EDTA; ER; OSTase; ProK; RM; RSA; TM; endoplasmic reticulum; ethylenediaminetetraacetic acid; glycosylation; membrane protein; oligosaccharyl transferase; protein topogenesis; proteinase K; rat serum albumin; rough microsomal membrane; translocation; translocon; transmembrane