N-Unsubstituted Glucosamine Residues in Heparan Sulfate and Their Potential Relation to Alzheimer's Disease

Heparan sulfate (HS) is a linear polysaccharide, located on the surface and in the extracellular matrix of most cells, that regulates functions of numerous proteins. HS-protein interaction is mainly mediated by sulfate groups found in N-sulfated (NS) regions of the HS, but may also involve rare HS substituents such as N-unsubstituted glucosamine (GlcNH2) residues. The location of GlcNH2 in an HS-epitope recognized by the monoclonal antibody 10E4, that specifically stains the prion lesions in scrapie-infected murine brain, suggests an involvement of GlcNH2 in prion disease and other amyloid-related disorders. HS in general is strongly associated with amyloidosis, including Alzheimer’s disease (AD). Therefore, the aims of this thesis were to structurally characterize GlcNH2-containing HS sequences found in native tissues, to further study HS epitopes recognized by 10E4, and to investigate the possible role(s) of GlcNH2 and other HS structures in binding to amyloid β peptide (Aβ) (core material in AD plaque lesions, also stained by 10E4). The GlcNH2 content (0.7-4% of total disaccharide units) varied between HS from different tissues. Most GlcNH2 units were found in poorly modified N-acetylated (NA-) or NA/NS-domains, located toward the polysaccharide-protein linkage region. Binding of human cerebral cortex HS to Aβ(1–40) monomers requires N-, 2- and 6-O-sulfation of HS, while binding to Aβ fibrils requires N- and 2-O-sulfation only. GlcNH2 units do not appreciably contribute to interaction with Aβ. Aβ fibril-binding HS domains also bind to fibroblast growth factor 2 (FGF-2), indicating that Aβ (neurotoxic) and FGF-2 (neuroprotective) may compete for common binding sites in HS. However, Aβ had no effect on FGF-2-induced MAPK signaling in NIH 3T3 fibroblasts. Continued studies on 10E4-antigenic HS epitope(s) showed that binding of 10E4 to the previously identified antigenic tetrasaccharide, ∆UA-GlcNH2-GlcA-GlcNAc, requires the nonreducing hexuronic acid (∆UA) to be 4,5 unsaturated (induced by lyase cleavage), and thus is artificial. Further studies are needed to clarify the potential involvement of GlcNH2 in 10E4-recognition of the native HS epitope(s)

Common binding sites for beta-amyloid fibrils and fibroblast growth factor-2 in heparan sulfate from human cerebral cortex

6 p.-4 fig.-1 tab.Heparan sulfate found in the cerebral plaques of Alzheimer's disease binds to beta-amyloid (Abeta) fibrils. This interaction has been proposed to enhance fibril deposition and mediate Abeta-induced glia activation and neurotoxicity. On the other hand, heparan sulfate augments signaling of fibroblast growth factor-2 (FGF-2), a neuroprotective factor that antagonizes the neurotoxic effects of Abeta. We defined structures in heparan sulfate from human cerebral cortex that bind Abeta fibrils. The minimal binding site is found in N-sulfated hexasaccharide domains and contains critical 2-O-sulfated iduronic acid residues. By contrast, binding of Abeta monomers requires, in addition, 6-O-sulfate groups on glucosamine residues. The binding specificity of fibrillar Abeta is shared by FGF-2, and we here show that cerebral heparan sulfate domains selected for binding to Abeta-(1-40) fibrils bind also to FGF-2. These data suggest that neurotoxic and neuroprotective signals may converge by competing for the same binding sites on the heparan sulfate chain.This work was supported by grants from the Swedish Medical Research Council (Grants K96–03P, 013004, and 2309), the Alzheimer Foundation, Sweden, Stiftelsen för Gamla Tjänarinnor, Thuréus Foundation, Svenska Lundbeckstiftelsen, Torsten och Ragnar Söderbergs Stiftelser, The Medical Faculty of Uppsala University, Polysackaridforskning AB (Uppsala, Sweden), and the program “Glycoconjugates in Biological Systems” sponsored by The Swedish Foundation for Strategic Research.Peer reviewe