The structural plasticity of heparan sulfate NA-domains and hence their role in mediating multivalent interactions is confirmed by high-accuracy 15N-NMR relaxation studies

Abstract

Considering the biological importance of heparan sulfate (HS) and the significant activity of its highly-sulfated regions (S-domains), the paucity of known functions for the non-sulfated NA-domains is somewhat puzzling. It has been suggested that chain dynamics within the NA-domains are the key to their functional role in HS. In this study, we investigate this hypothesis using state-of-the-art nuclear magnetic resonance (NMR) experiments at multiple frequencies. To resolve the problem of severe overlap in 1H-NMR spectra of repetitive polysaccharides from proteoglycans, we have prepared oligosaccharides with the chemical structure of HS NA-domains containing the 15N nucleus, which has enough chemical shift dispersion to probe the central residues of octasaccharides at atomic resolution using 600 MHz NMR. By performing NMR relaxation experiments at three magnetic-field strengths, high quality data on internal dynamics and rotational diffusion was obtained. Furthermore, translational diffusion could also be measured by NMR using pulse field gradients. These experimental data were used, in concert with molecular dynamics simulations, to provide information on local molecular shape, greatly aiding our relaxation analyses. Our results, which are more accurate than those presented previously, confirm the higher flexibility of the NA-domains as compared with reported data on S-domains. It is proposed that this flexibility has two functional roles. First, it confers a greater area of interaction from the anchoring point on the core protein for the bioactive S-domains. Secondly, it allows multiple interactions along the same HS chain that are dynamically independent of each other.Electronic Supplementary MaterialThe online version of this article (doi:10.1007/s10719-007-9081-9) contains supplementary material, which is available to authorized users.