Quantification of Amyloid Precursor Protein Isoforms Using Quantification Concatamer Internal Standard

• Published in: Analytical chemistry (Washington) 2013-01, Vol.85 (1), p.303-307
• Main Authors: Chen, Junjun, Wang, Meiyao, Turko, Illarion V
• Format: Article
• Language: English
• Subjects: Alzheimer Disease - metabolism; Alzheimer Disease - pathology; Alzheimer's disease; Amino Acid Sequence; Amino acids; Amyloid beta-Protein Precursor - analysis; Amyloid beta-Protein Precursor - metabolism; Amyloid beta-Protein Precursor - standards; Carbon Isotopes - chemistry; Cerebral Cortex - metabolism; Chromatography, High Pressure Liquid; Gene expression; Humans; Isotope Labeling; Molecular Sequence Data; Nitrogen Isotopes - chemistry; Pathogenesis; Peptides; Peptides - analysis; Peptides - standards; Protein Isoforms - analysis; Protein Isoforms - metabolism; Protein Isoforms - standards; Proteins; Reference Standards; Tandem Mass Spectrometry - standards; Trypsin - metabolism
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac3033239

It is likely that expression and/or post-translational generation of various protein isoforms can be indicative of initial pathological changes or pathology development. However, selective quantification of individual protein isoforms remains a challenge, because they simultaneously possess common and unique amino acid sequences. Quantification concatamer (QconCAT) internal standards were originally designed for a large-scale proteome quantification and are artificial proteins that are concatamers of tryptic peptides for several proteins. We developed a QconCAT for quantification of various isoforms of amyloid precursor protein (APP). APP-QconCAT includes tryptic peptides that are common for all isoforms of APP concatenated with those tryptic peptides that are unique for specific APP isoforms. Isotope-labeled APP-QconCAT was expressed, purified, characterized, and further used for quantification of total APP, APP695, and amyloid-β (Aβ) in the human frontal cortex from control and severe Alzheimer’s disease donors. Potential biological implications of our quantitative measurements are discussed. It is also expected that using APP-QconCAT(s) will advance our understanding of biological mechanism by which various APP isoforms involved in the pathogenesis of Alzheimer’s disease.