Beta-amyloid precursor protein of Alzheimer disease occurs as 110- to 135-kilodalton membrane-associated proteins in neural and nonneural tissues.

Abstract

Progressive cerebral deposition of extracellu- (15,16). A related issue concerns the mechanism whereby the normal, age-related deposition of (AP in selected brain regions and vessels is augmented in AD. The local proteolytic processing of the precursor and the role of amyloidassociated proteins, such as the serine protease inhibitor a1-antichymotrypsin (24), require elucidation.Although molecular studies of 8APP have advanced rapidly, the native precursor protein in human brain and other tissues has not been detected and characterized. To begin to address some of the questions posed above, we produced antibodies to synthetic peptides with sequences predicted from amyloidogenic and nonamyloidogenic regions of J3APP and have identified forms of the precursor molecule in brain, nonneural tissues, and cultured cells of several species. METHODSPreparation of Antibodies to Synthetic Peptides. Several peptides were synthesized according to the sequence deduced from a P3APP cDNA (14). Those used in this study were: amino acid residues 597-624 (peptide (31-28), comprising the first 28 residues of PAP; residues 676-695 (peptide C1), comprising the 20 C-terminal amino acids of PAPP; and residues 681-695, a second C-terminal peptide (C2), comprising the last 15 residues. The latter peptide and an antiserum to it were the gifts of Tsuyoshi Ishii (Psychiatric Research Institute, Tokyo). Peptide C1 was HPLC-purified and coupled to edestin. Peptide C2 had been coupled to keyhole limpet hemocyanin. Peptide p1-28 was injected uncoupled. Antisera were assayed by serial dilutions on dot blots of unconjugated peptide. Previously characterized antibodies used in this study included two antisera (A and C) to the -4-kDa P3AP purified from amyloid-rich fractions of AD cortex (12,25), two antisera (Fl and Ph) to the -4-kDa ,8AP purified from AD meningovascular amyloid (10), a paired helical filament-specific antiserum having no reaction with amyloid deposits (26), and an antiserum to heat-stable microtubule-associated proteins (principally X and MAP 2) purified from fetal human brain (25).Abbreviations: AD, Alzheimer disease; flAP, f8-amyloid protein; f3APP, 13-amyloid precursor polypeptide. 7341The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Gelatinase A possesses a β‐secretase‐like activity in cleaving the amyloid protein precursor of Alzheimer's disease

“…However, at 100 ng/ml gelatinase A, a 100-110 kDa band was seen (lane 4). This band was approximately 10 kDa lower in molecular mass than the major cellular form of APP [8,15,16]. At the highest concentration of gelatinase A (1 pg/ml), a second cleavage product of 70 kDa was detected (lane 5).…”

“…q heir studies showed that a synthetic peptide homologous to r,~sidues [10][11][12][13][14][15][16][17][18][19][20] residue known to be the site of cleavage of the ~-secretase. However, as gelatinase A was activated by a procedure employing trypsin digestion and stromelysin activation, it was not conclusively demonstrated in that study that the activated gelatinase A was free of trypsin activity.…”

An Etiological Role of Amyloidogenic Carboxyl‐Terminal Fragments of the β‐Amyloid Precursor Protein in Alzheimer's Disease

“…Immunoblots of the fractionated proteins from the transgenic mouse brains showed the reactivity with antibody 369A to the CT 50 amino acids of /IAPP (Buxbaum et al, 1990) and with C4, an antibody to the CT 10 amino acids of /IAPP (Selkoe et al, 1988).…”

“…The recent concentration on other potentially amyloidogenic products of /IAPP has produced interesting candidates, the most promising of which are the amyloidogenic carboxyl-terminal (CT) fragments of /IAPP. The transgenic mice expressing /IAPP and presenilins were not, however, examined for the presence of CT 100.First, CT peptides have been found not only in various cultured cells (Maruyama et al, 1990;Wolf et al, 1990;Dyrks et al, 1992;Estus et al, 1992; Gandy et al., 1992a,b;Golde et al, 1992;Haass et al, 1992b) but also in paired helical filaments (Caputo et al, 1992), in senile plaques (Selkoe et al, 1988), in microvessels (Tamaoka et al, 1992), and in choroid plexus from human brain in AD, in the white matter of Down's syndrome brains (Tokuda et al, 1995), and in human platelets (Tables 1 and 2). CT fragments with molecular masses of 12-16 kDa have also been found in media and cytosol of lymphoblastoid cells obtained from patients with earlyor late-onset FAD (Matsumoto, 1994) and Down's syndrome (Kametani et al, 1994).…”

Molecular Cloning of Human cDNA with a Sequence Highly Similar to that of the Dihydrofolate Reductase Gene in Brain Libraries Derived from Alzheimer's Disease Patients

“…This suggests that an abnormality in APP itself and, factor(s) that modifies APP processing, are sufficient for the production of senile plaque, which requires a long period of maturation. In Alzheimer's disease brain, in which an abnormal APP processing pathway that accumulates selfaggregatable PA4-harbouring peptides predominates, amyloid in a P-pleated sheet conformation is induced under physiological conditions (Selkoe et al, 1988;Estus et al, 1992) whereas, in normal brain, intra-PA4 proteolysis by an APP secretase precludes p-amyloid formation (Esch et al, 1990;McDermott and Gibson, 1991).…”