title: Integration of transmembrane domains is regulated by their downstream sequences.
abstract: The Sec61 translocon catalyzes translocation of proteins into the endoplasmic reticulum and the lateral integration of transmembrane segments into the lipid bilayer. Integration is mediated by the hydrophobicity of a polypeptide segment consistent with thermodynamic equilibration between the translocon and the lipid membrane. Integration efficiency of a generic series of increasingly hydrophobic sequences (H-segments) was found to diverge significantly in different reporter constructs as a function of the ∼100 residues that are C-terminal to the H-segments. The hydrophobicity threshold of integration was considerably lowered through insertion of generic ∼20-residue peptides either made of flexible glycine-serine repeats, containing multiple negative charges, or consisting of an oligoproline stretch. A highly flexible, 100-residue glycine-serine stretch maximally enhanced this effect. The apparent free energy of integration was found to be changed by more than 3 kcal/mol with the downstream sequences tested. The C-terminal sequences could also be shown to affect integration of natural mildly hydrophobic sequences. The results suggest that the conformation of the nascent polypeptide in the protected cavity between the ribosome and translocon considerably influences the release of the H-segment into the bilayer.
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 70, Basel CH-4056, Switzerland.
authors.forename: Tina
authors.initials: T
authors.lastname: Junne
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 70, Basel CH-4056, Switzerland martin.spiess@unibas.ch.
authors.forename: Martin
authors.identifier: 0000-0001-7139-0550
authors.initials: M
authors.lastname: Spiess
mesh_terms: D000595:Amino Acid Sequence; D003062:Codon / Q000235:genetics; D017930:Genes, Reporter; D057927:Hydrophobic and Hydrophilic Interactions; D008565:Membrane Proteins / Q000737:chemistry*; D008958:Models, Molecular; D011392:Proline / Q000737:chemistry; D000072417:Protein Domains; D013329:Structure-Activity Relationship
chemical_list: D003062:Codon; D008565:Membrane Proteins; D011392:Proline
keywords: Endoplasmic reticulum; Membrane insertion; Protein translocation; Sec61; Translocon