Analysis of the primary and secondary structure of the glycosaminoglycans from alternative sources (natural or synthetic)

The PhD project developed during these last three years concerned the analysis of the primary and secondary structure of Glycosamminoglycans. The aim of the work was the collection of new information (spectroscopic and conformational ones) about Hyaluronic acid, Chondroitin sulfate and Keratan sulfate, in order to amplify the literature data available about their structures-function relationship. Glycosaminoglycans (GAGs), are a family of linear polydisperse polysaccharides that take part in several physiological phenomena, including neuronal development, cell–matrix interactions, and activation of chemokines, enzymes, and growth factors. These chains are typically linked to a protein core forming proteoglycans at the cell surface or in the extracellular matrix. The ability of GAGs to regulate these processes is attributed to their complex structure, which arises from extensive modifications (sulfation, deacetylation and epimerization) of a nonsulfated precursor consisting of hexosamine (D-Glucosamine, D-Galactosamine) and either hexuronic acid (D-Glucuronic acid, L-iduronic acid) or galactose residues, arranged in an alternating linear sequence. Based on of the chemical structure backbone, there are four classes of GAGs: Hyaluronan (HA), Heparin (HP)/Heparan sulfate (HS), Chondroitin (CS)/Dermatan sulfate (DS) and Keratan sulfate (KS). Despite their simple backbone sequence, glycosaminoglycans are complex molecules characterized by a heterogeneous structure, for this reason the investigation on them is really challenging and it is performed usually combining many techniques. In this work, analyses were carried out applying a combination of chemical, enzymatic and spectroscopic tools. In some cases the spectroscopic data collected were analyzed with the support of theoretical data obtained through molecular mechanics and dynamics simulations. The complete work, for a easier organization, is divided into two parts: • Structural characterization of natural GAG samples:  Chondroitin sulfate from the cartilage of Scyliorhinus canicula, Raja brachyura, and Torpedo nobiliana and also from the skin of Raja brachyura.  Keratan sulfate from the cartilage of the Scyliorhinus canicula. • Conformational analyses of GAG oligosaccharides:  Decasaccharide of hyaluronic acid  Hexasaccharide of unsulfate chondroitin  Dodedecasaccharide of chondroitin sulfate  Pentasaccharides of keratan sulfate