Mass spectrometry of purified amyloid beta protein in Alzheimer's disease.
The amyloid beta-protein (A beta) that is progressively deposited in Alzheimer's disease (AD) arises from proteolysis of the integral membrane protein, beta-amyloid precursor protein (beta APP). Although A beta formation appears to play a seminal role in AD, only a few studies have examined the chemical structure of A beta purified from brain, and there are discrepancies among the findings. We describe a new method for the rapid extraction and purification of A beta that minimizes artifactual proteolysis. A beta purified by two-dimensional reverse-phase HPLC was analyzed by combined amino acid sequencing and mass spectrometry after digestion with a lysylendopeptidase. The major A beta peptide in the cerebral cortex of all five AD brains examined was aspartic acid 1 to valine 40. A minor species beginning at glutamic acid 3 but blocked by conversion to pyroglutamate was also found in all cases. A species ending at threonine 43 was detected, varying from approximately 5 to 25% of total A beta COOH-terminal fragments. Peptides ending with valine 39, isoleucine 41, or alanine 42 were not detected, except for one brain with a minor peptide ending at valine 39. Our findings suggest that A beta 1-40 is the major species of beta-protein in AD cerebral cortex. A beta 1-40 and A beta 1-43 peptides could arise independently from beta APP, or A beta 1-43 could be the initial excised fragment, followed by digestion to yield A beta 1-40. These analyses of native A beta in AD brain recommend the use of synthetic A beta 1-40 peptide to model amyloid fibrillogenesis and toxicity in vitro.
Mori
H
H
Department of Neuropathology, University of Tokyo, Japan.
Takio
K
K
Ogawara
M
M
Selkoe
D J
DJ
D000544:Alzheimer Disease / Q000378:metabolism* / Q000473:pathology; D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000737:chemistry* / Q000302:isolation & purification; D002540:Cerebral Cortex / Q000737:chemistry / Q000473:pathology; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000302:isolation & purification; D012697:Serine Endopeptidases
D016229:Amyloid beta-Peptides; D010446:Peptide Fragments; D012697:Serine Endopeptidases; C036813:lysyl endopeptidase