Inhaltszusammenfassung

The secretory N-terminal domain of the Alzheimer amyloid precursor protein (sAPP) evokes specific responses in cells on binding to their surfaces. Because APP is expressed in a large variety of cell types, the localization of sAPP binding requires detection techniques that selectively recognize sAPP as a ligand. For this purpose, we prepared antibodies against recombinant sAPP695 (sAPPrec) previously expressed in E. coli. Such antibodies were found to distinguish between sAPPrec and cellular ...The secretory N-terminal domain of the Alzheimer amyloid precursor protein (sAPP) evokes specific responses in cells on binding to their surfaces. Because APP is expressed in a large variety of cell types, the localization of sAPP binding requires detection techniques that selectively recognize sAPP as a ligand. For this purpose, we prepared antibodies against recombinant sAPP695 (sAPPrec) previously expressed in E. coli. Such antibodies were found to distinguish between sAPPrec and cellular APP or sAPP, as shown by immunocytochemistry and by immunoblot. in addition, they allowed the selective localization of bound sAPPrec on cell surfaces without any signal from cellular APP or sAPP. Saturation of sAPPrec binding to cell surfaces, as determined radiometrically, was reached at 10 nM [I-125]-sAPPrec. Binding was specific because it was almost completely inhibited by a 100-fold excess of unlabeled sAPPrec. This specificity of binding was confirmed by surface plasmon resonance spectroscopy. Binding of sAPPrec to cell surfaces occurred in patches and was dependent on the state of cell differentiation. The sAPPrec used in this study contains heparin binding sites, but enzymatic removal of cell surface associated heparin did not affect sAPPrec binding. Aldehyde fixation of cells strongly inhibited their ability to bind sAPPrec. The data point to a fixation-sensitive sAPPrec binding protein which is detectable in the form of patches and therefore is part of assembled cell surface microdomains.» weiterlesen» einklappen

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DDC Sachgruppe:
Medizin