title: Transmembrane orientation of signal-anchor proteins is affected by the folding state but not the size of the N-terminal domain.
abstract: Upon insertion of a signal-anchor protein into the endoplasmic reticulum membrane, either the C-terminal or the N-terminal domain is translocated across the membrane. Charged residues flanking the transmembrane domain are important determinants for this decision, but are not necessarily sufficient to generate a unique topology. Using a model protein that is inserted into the membrane to an equal extent in either orientation, we have tested the influence of the size and the folding state of the N-terminal domain on the insertion process. A small zinc finger domain or the full coding sequence of dihydrofolate reductase were fused to the N-terminus. These stably folding domains hindered or even prevented their translocation. Disruption of their structure by destabilizing mutations largely restored transport across the membrane. Translocation efficiency, however, did not depend on the size of the N-terminal domain within a range of 40-237 amino acids. The folding behavior of the N-terminal domain is thus an important factor in the topogenesis of signal-anchor proteins.
authors.affiliation: Department of Biochemistry, Biozentrum, University of Basel, Switzerland.
authors.forename: A J
authors.initials: AJ
authors.lastname: Denzer
authors.forename: C E
authors.initials: CE
authors.lastname: Nabholz
authors.forename: M
authors.initials: M
authors.lastname: Spiess
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins / Q000378:metabolism; D001483:Base Sequence; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000737:chemistry; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011487:Protein Conformation; D017510:Protein Folding; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D013762:Tetrahydrofolate Dehydrogenase / Q000737:chemistry / Q000235:genetics; D016335:Zinc Fingers / Q000235:genetics
chemical_list: D037263:Asialoglycoprotein Receptor; D001212:Asialoglycoproteins; D017931:DNA Primers; D008565:Membrane Proteins; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins; D013762:Tetrahydrofolate Dehydrogenase