Binding studies of truncated variants of the Aβ peptide to the V-domain of the RAGE receptor reveal Aβ residues responsible for binding

Alzheimer's disease (AD) symptoms correlate with the concentration of soluble, although not necessarily monomeric forms of A beta peptide in the brain parenchyma. The RAGE receptor has been implicated as the protein responsible for active transport of A beta from blood circulation to the brain. In murine models of AD, inhibition of the A beta:RAGE interaction decreases the levels of A beta in the brain. Inhibition of the A beta:RAGE interaction would be a promising alternative for the therapy of AD. Rational design of an A beta:RAGE interaction blocker requires detailed knowledge of the structure of the complex. However, the binding domain of RAGE is natively unfolded in physiological conditions, which severely hampers the application of classic methods of protein structure analysis to the design of an antagonist. Here, alternative methods are used to characterize the structural properties of the RAGE-ligand binding domain and to monitor the binding of a series of truncated variants of A beta. Using intrinsic RAGE tryptophan fluorescence and mass spectrometry of non-covalent protein-ligand complexes we have identified shorter versions of A beta that bind to the RAGE V-domain. We have also shown in cell culture experiments that a selected shortened version of A beta effectively inhibits full-length A beta RAGE-mediated, cell uptake. Thus, a truncated version of A beta capable of blocking its receptor-mediated internalization was established, revealing the binding code and providing the lead compound in the process of drug design. (C) 2011 Elsevier B.V. All rights reserved.