Functional implications of the presenilin dimerization: reconstitution of gamma-secretase activity by assembly of a catalytic site at the dimer interface of two catalytically inactive presenilins.
Presenilins are the catalytic components of gamma-secretase, an intramembrane-cleaving protease whose substrates include beta-amyloid precursor protein (betaAPP) and the Notch receptors. These type I transmembrane proteins undergo two distinct presenilin-dependent cleavages within the transmembrane region, which result in the production of Abeta and APP intracellular domain (from betaAPP) and the Notch intracellular domain signaling peptide. Most cases of familial Alzheimer's disease are caused by presenilin mutations, which are scattered throughout the coding sequence. Although the underlying molecular mechanism is not yet known, the familial Alzheimer's disease mutations produce a shift in the ratio of the long and short forms of the Abeta peptide generated by the gamma-secretase. We and others have previously shown that presenilin homodimerizes and suggested that a presenilin dimer is at the catalytic core of gamma-secretase. Here, we demonstrate that presenilin transmembrane domains contribute to the formation of the dimer. In-frame substitution of the hydrophilic loop 1, located between transmembranes I and II, which modulates the interactions within the N-terminal fragment/N-terminal fragment dimer, abolishes both presenilinase and gamma-secretase activities. In addition, by reconstituting gamma-secretase activity from two catalytically inactive presenilin aspartic mutants, we provide evidence of an active diaspartyl group assembled at the interface between two presenilin monomers. Under our conditions, this catalytic group mediates the generation of APP intracellular domain and Abeta but not Notch intracellular domain, therefore suggesting that specific diaspartyl groups within the presenilin catalytic core of gamma-secretase mediate the cleavage of different substrates.
Cervantes
Sara
S
Departament de Genètica, Facultat de Biologia, Universitat de Barcelona, Avda. Diagonal 645, E-08028 Barcelona, Spain.
Saura
Carlos A
CA
Pomares
Esther
E
Gonzàlez-Duarte
Roser
R
Marfany
Gemma
G
D053829:Amyloid Precursor Protein Secretases; D000818:Animals; D001224:Aspartic Acid / Q000737:chemistry; D016282:Aspartic Acid Endopeptidases; D002384:Catalysis; D020134:Catalytic Domain; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D019281:Dimerization; D004330:Drosophila; D029721:Drosophila Proteins; D010450:Endopeptidases / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D015151:Immunoblotting; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D053764:Presenilin-1; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D051880:Receptors, Notch; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D014162:Transfection; D025801:Ubiquitin / Q000737:chemistry / Q000378:metabolism
D029721:Drosophila Proteins; D008565:Membrane Proteins; C497759:N protein, Drosophila; C508853:PSEN1 protein, human; D010455:Peptides; D053764:Presenilin-1; D051880:Receptors, Notch; D011993:Recombinant Fusion Proteins; D025801:Ubiquitin; D001224:Aspartic Acid; D004247:DNA; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human