ALZHEIMER AMYLOID BETA/A4 PEPTIDE BINDING-SITES AND A POSSIBLE APP-SECRETASE ACTIVITY ASSOCIATED WITH RAT-BRAIN CORTICAL MEMBRANES

We carried out ligand binding experiments on membranes from rat brain cortical grey matter using radioiodinated beta/A4 8-17, with non-specific binding determined by the addition of 10-mu-M unlabelled peptide. Specific, reversible binding amounted to 60-75% of total binding and showed a clear dependence on time, temperature, pH and membrane concentration. Kinetic analyses indicated a high-affinity binding site with an apparent K(D) of 440 pM. However, the ligand was partly degraded with loss of the Ser8, Lys16 and Leu17 residues. Excision of the two C-terminal amino acids was inhibited by EDTA, EGTA, dithiothreitol or Zn2+ but was stimulated by Ca2+ or Mn2+. These studies demonstrate high-affinity binding sites for beta/A4 8-17 (or its derivatives) in rat brain, suggesting that this region may contain a physiologically important amino acid sequence and identify a potential membrane-associated amyloid precursor protein (APP) secretase activity.