title: Functional domains in presenilin 1: the Tyr-288 residue controls gamma-secretase activity and endoproteolysis.
abstract: Processing of the Alzheimer amyloid precursor protein (APP) into the amyloid beta-protein and the APP intracellular domain is a proteolysis event mediated by the gamma-secretase complex where presenilin (PS) proteins are key constituents. PS is subjected to an endoproteolytic cleavage, generating a stable heterodimer composed of an N-terminal and a C-terminal fragment. Here we aimed at further understanding the role of PS in endoproteolysis, in proteolytic processing of APP and Notch, and in assembly of the gamma-secretase complex. By using a truncation protocol and alanine scanning, we identified Tyr-288 in the PS1 N-terminal fragment as critical for PS-dependent intramembrane proteolysis. Further mutagenesis of the 288 site identified mutants differentially affecting endoproteolysis and gamma-secretase activity. The Y288F mutant was endoproteolyzed to the same extent as wild type PS but increased the amyloid beta-protein 42/40 ratio by approximately 75%. In contrast, the Y288N mutant was also endoproteolytically processed but was inactive in reconstituting gamma-secretase in PS null cells. The Y288D mutant was deficient in both endoproteolysis and gamma-secretase activity. All three mutant PS1 molecules were incorporated into gamma-secretase complexes and stabilized Pen-2 in PS null cells. Thus, mutations at Tyr-288 do not affect gamma-secretase complex assembly but can differentially control endoproteolysis and gamma-secretase activity.
authors.affiliation: Department of Neurotec, Division of Experimental Geriatrics, Karolinska Institutet, Novum, SE-141 86 Huddinge, Sweden.
authors.forename: Hanna
authors.initials: H
authors.lastname: Laudon
authors.forename: Helena
authors.initials: H
authors.lastname: Karlström
authors.forename: Paul M
authors.initials: PM
authors.lastname: Mathews
authors.forename: Mark R
authors.initials: MR
authors.lastname: Farmery
authors.forename: Samuel E
authors.initials: SE
authors.lastname: Gandy
authors.forename: Johan
authors.initials: J
authors.lastname: Lundkvist
authors.forename: Urban
authors.initials: U
authors.lastname: Lendahl
authors.forename: Jan
authors.initials: J
authors.lastname: Näslund
mesh_terms: D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases; D000818:Animals; D016282:Aspartic Acid Endopeptidases; D002478:Cells, Cultured; D004622:Embryo, Mammalian / Q000166:cytology; D010450:Endopeptidases / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D017930:Genes, Reporter; D015151:Immunoblotting; D008156:Luciferases / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry*; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011233:Precipitin Tests; D053764:Presenilin-1; D017434:Protein Structure, Tertiary; D013234:Stem Cells / Q000166:cytology; D013347:Subcellular Fractions; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry*
chemical_list: D008565:Membrane Proteins; D053764:Presenilin-1; D014443:Tyrosine; D008156:Luciferases; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509282:Bace1 protein, mouse; D000409:Alanine