title: A nine-transmembrane domain topology for presenilin 1.
abstract: Presenilin (PS) provides the catalytic core of the gamma-secretase complex. Gamma-secretase activity leads to generation of the amyloid beta-peptide, a key event implicated in the pathogenesis of Alzheimer disease. PS has ten hydrophobic regions, which can all theoretically form membrane-spanning domains. Various topology models have been proposed, and the prevalent view holds that PS has an eight-transmembrane (TM) domain organization; however, the precise topology has not been unequivocally determined. Previous topological studies are based on non-functional truncated variants of PS proteins fused to reporter domains, or immunocytochemical staining. In this study, we used a more subtle N-linked glycosylation scanning approach, which allowed us to assess the topology of functional PS1 molecules. Glycosylation acceptor sequences were introduced into full-length human PS1, and the results showed that the first hydrophilic loop is oriented toward the lumen of the endoplasmic reticulum, whereas the N terminus and large hydrophilic loop are in the cytosol. Although this is in accordance with most current models, our data unexpectedly revealed that the C terminus localized to the luminal side of the endoplasmic reticulum. Additional studies on the glycosylation pattern after TM domain deletions, combined with computer-based TM protein topology predictions and biotinylation assays of different PS1 mutants, led us to conclude that PS1 has nine TM domains and that the C terminus locates to the lumen/extracellular space.
authors.affiliation: Department of Neurotec, Division of Experimental Geriatrics, Karolinska Institutet, Novum, SE-141 86 Huddinge, Sweden. Hanna.Laudon@neurotec.ki.se
authors.forename: Hanna
authors.initials: H
authors.lastname: Laudon
authors.forename: Emil M
authors.initials: EM
authors.lastname: Hansson
authors.forename: Karin
authors.initials: K
authors.lastname: Melén
authors.forename: Anna
authors.initials: A
authors.lastname: Bergman
authors.forename: Mark R
authors.initials: MR
authors.lastname: Farmery
authors.forename: Bengt
authors.initials: B
authors.lastname: Winblad
authors.forename: Urban
authors.initials: U
authors.lastname: Lendahl
authors.forename: Gunnar
authors.initials: G
authors.lastname: von Heijne
authors.forename: Jan
authors.initials: J
authors.lastname: Näslund
mesh_terms: D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D001665:Binding Sites; D019921:Biotinylation; D002462:Cell Membrane / Q000378:metabolism*; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D010450:Endopeptidases / Q000737:chemistry; D004721:Endoplasmic Reticulum / Q000378:metabolism; D005109:Extracellular Matrix / Q000378:metabolism; D017353:Gene Deletion; D017930:Genes, Reporter; D006026:Glycoside Hydrolases / Q000494:pharmacology; D006031:Glycosylation; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D008156:Luciferases / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D053764:Presenilin-1; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D019809:Streptavidin / Q000737:chemistry
chemical_list: D008565:Membrane Proteins; C508853:PSEN1 protein, human; D053764:Presenilin-1; D019809:Streptavidin; D008156:Luciferases; D006026:Glycoside Hydrolases; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human; C509282:Bace1 protein, mouse