title: The topogenic contribution of uncharged amino acids on signal sequence orientation in the endoplasmic reticulum.
abstract: Signal sequences for insertion of proteins into the endoplasmic reticulum induce translocation of either the C- or the N-terminal sequence across the membrane. The end that is translocated is primarily determined by the flanking charges and the hydrophobic domain of the signal. To characterize the hydrophobic contribution to topogenesis, we have challenged the translocation machinery in vivo in transfected COS cells with model proteins differing exclusively in the apolar segment of the signal. Homo-oligomers of hydrophobic amino acids as different in size and shape as Val(19), Trp(19), and Tyr(22) generated functional signal sequences with similar topologies in the membrane. The longer a homo-oligomeric sequence of a given residue, the more N-terminal translocation was obtained. To determine the topogenic contribution of all uncharged amino acids in the context of a hydrophobic signal sequence, two residues in a generic oligoleucine signal were exchanged for all uncharged amino acids. The resulting scale resembles a hydrophobicity scale with the more hydrophobic residues promoting N-terminal translocation. In addition, the helix breakers glycine and proline showed a position-dependent effect, which raises the possibility of a conformational contribution to topogenesis.
authors.affiliation: Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 Basel, Switzerland.
authors.forename: K
authors.initials: K
authors.lastname: Rösch
authors.forename: D
authors.initials: D
authors.lastname: Naeher
authors.forename: V
authors.initials: V
authors.lastname: Laird
authors.forename: V
authors.initials: V
authors.lastname: Goder
authors.forename: M
authors.initials: M
authors.lastname: Spiess
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D037263:Asialoglycoprotein Receptor; D001483:Base Sequence; D019556:COS Cells; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D007425:Intracellular Membranes / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D021382:Protein Sorting Signals / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D014162:Transfection; D014364:Tryptophan; D014443:Tyrosine; D014633:Valine
chemical_list: D037263:Asialoglycoprotein Receptor; D009838:Oligodeoxyribonucleotides; D021382:Protein Sorting Signals; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D014443:Tyrosine; D014364:Tryptophan; D014633:Valine