Changes in heparan sulfate structure and location have long been associated with Alzheimer’s disease, but the functional significance of this has remained elusive. Recently, it has been demonstrated that heparan sulfate drives taupathy type Alzheimer’s disease in a variety of model systems, the foundation of the EC-FET-OPEN programme, ArrestAD. ArrestAD is restricted to work on human samples, despite the far greater tractability of model systems. This project will, therefore, work on mouse brain, derived from wild-type animals and from transgenic animals with taupathy. The latter tissue will be supplied by the Coordinator of ArrestAD. The first phase of the project will use a proteomics approach to identify heparin-binding proteins in normal mouse brain and in brain from taupathy mice. The streamlined heparin-affinity proteomics pipeline derived in a previous project on pancreas will be used. Proteins from plasma membrane/extracellular matrix enriched fractions will be subjected to heparin affinity chromatography and heparin-binding proteins quantified by label-free mass spectrometry by the CPR. The functional relationships of these proteins will be analysed using conventional systems biology tools to which we have access, e.g., IPA. There are two possible second phases of the project, depending on the interpretation of the results of the first phase. (1) Identification of lysine residues responsible for heparin binding in brain proteins. Our existing selective labelling method, used with individual pure proteins, will be adapted to a complex mixture, initially using simple mixtures of fibroblast growth factors with well-characterised heparin-binding sites. From there, measurements will be made on the heparin-binding proteins from mouse brain. (2) Organisation of matrix of fixed tissue from normal and taupathy. Here, recombinant Halotag-fibroblast growth factor fusion proteins would be used as probes for the spatial disposition of their heparan sulfate binding sites. The work would then progress to measure the dynamics of the fibroblast growth factor using fluorescence recovery after photobleaching to identify key differences in the supramolecular organisation of matrix in the normal and diseased tissue.