Studies of carbohydrate structure, properties and interactions by nmr spectroscopy

Abstract: In this thesis the structure, properties and interactions of different types of carbohydrates were investigated by NMR spectroscopy. The structures of kappa- (κ) and kappa/mu- (κ/μ) carrageenan oligosaccharides were analyzed by studying the hydroxy protons. It was shown that a hydrogen bonding interaction is present across the 1-4 glycosidic linkages of μ-carrabiose in the κμκ hexa- and κμμκ octasaccharides. The occurrence of hydrogen bonding in κ/μ-carrageenan oligosaccharides may suggest that μ-carrabiose units, mostly found in the non-helicoidal regions of κ-carrageenans have an underestimated role in the structural organization of the κ-carrageenan gel network. Hydroxy proton NMR was also used to study the effect of trehalose on the hydration and hydrogen bonding in lactose in aqueous solutions. The small effects of trehalose on the hydration and hydrogen bonding interaction in lactose were very similar to those found for sucrose. The results suggested that, at concentrations below 40% (w/w), it is the concentration of hydroxy groups that governs sugar-sugar and sugar-water interactions rather than the type of sugar. A method using diffusion-edited NMR spectroscopy was developed for solvent suppression when determining the mannuronic (M) to guluronic (M) acid ratio in alginate polysaccharides. The method could be employed to determine the M/G-ratio at temperatures below 50 °C. Through all of the work in the thesis diffusion-edited NMR experiments also proved to be practical for studies of biomolecules, to for example selectively remove interfering signals from buffer to enable the interpretation of sample signals. The activities and specificities of four different glycosaminoglycan (GAG) sulfatases from the human gut commensal Bacteroides thetaiotaomicron were determined. One of the sulfatases, BT3349, was found to be the first bacterial GAG endolytic-O-sulfatase, with chondroitin specific GalNAc-4-O-sulfatase activity. The other three enzymes were shown to possess strictly exolytic activity, BT3333 as a GalNAc-6-O-sulfatase, BT4656 as a GlcNAc-6-O-sulfatase and the third one, BT1596, as a Δ-4-hexuronate-2-O-sulfatase.