Bacterial and cell-free production of APP671-726 containing amyloid precursor protein transmembrane and metal-binding domains

• Published in: Biochemistry (Moscow) 2013-11, Vol.78 (11), p.1263-1271
• Main Authors: Bocharova, O. V., Urban, A. S., Nadezhdin, K. D., Bocharov, E. V., Arseniev, A. S.
• Format: Article
• Language: English
• Subjects: Alzheimer Disease - metabolism; Alzheimer Disease - pathology; Alzheimer's disease; Amino Acid Sequence; Amyloid beta-Protein Precursor - chemistry; Amyloid beta-Protein Precursor - genetics; Amyloid beta-Protein Precursor - metabolism; Amyloid Precursor Protein Secretases - metabolism; Bacteria; Bacteriology; Biochemistry; Biomedical and Life Sciences; Biomedicine; Bioorganic Chemistry; Biosynthesis; Dimerization; Enzymes; Escherichia coli - metabolism; Gene mutations; Genetic aspects; Glycoproteins; Humans; Isotope Labeling; Life Sciences; Lipid Bilayers - chemistry; Lipid Bilayers - metabolism; Microbiology; Molecular Sequence Data; Mutation; Nuclear Magnetic Resonance, Biomolecular; Peptide Fragments - biosynthesis; Peptide Fragments - chemistry; Peptide Fragments - isolation & purification; Peptides; Physiological aspects; Protein Structure, Tertiary; Proteins; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Recombinant Proteins - isolation & purification; Risk factors
• ISSN: 0006-2979; 1608-3040
• DOI: 10.1134/S0006297913110060

More than half of the mutations associated with familiar Alzheimer’s disease have been found in the transmembrane domain of amyloid precursor protein (APP). These pathogenic mutations presumably influence the APP transmembrane domain structural and dynamic properties and result in its conformational change or/and lateral dimerization. Despite much data about the pathogenesis of Alzheimer’s disease, the initial steps of the pathogenesis remain unclear so far. For the investigation of the molecular basis of Alzheimer’s disease, we selected amyloid precursor protein fragment APP671-726 containing the transmembrane and metal-binding domains. This fragment is the substrate of the γ-secretase complex whose abnormal activity leads to the formation of amyloidogenic Aβ42 peptides. This work for the first time describes a highly effective cell-free APP671-726 production method and improved method of bacterial synthesis. Both methods yield milligram quantities of isotope-labeled protein for structural study by high resolution NMR spectroscopy in membrane mimicking milieus.