1H NMR studies of molecular interactions of carbohydrates in aqueous solutions

摘要

The formation and structure of inclusion complexes between α-cyclodextrin (α-CD) and adamantane, 1-adamantanol, 1-(hydroxymethyl)-adamantane, 2-adamantanol and 1,3-adamantanediol in aqueous solutions were studied by 1H NMR spectroscopy using both exchangeable and non-exchangeable protons. Complexes were formed with all adamantane derivatives with the exception of 1,3-adamantanediol. Similarities between the α-CD/adamantane and the α-CD/2-adamantanol complexes were evidenced by the appearance of a narrow and upfield shifted O(3)H signal for α-CD in the complex. α-CD formed 1:1 complexes with 1-adamantanol and 1-(hydroxymethyl)-adamantane. In both cases the O(2)H signal of α-CD was broadened with higher concentration of guest molecule and low temperatures. 1H NMR studies of the hydrogen bonding network in mono-altro-β-cyclodextrin and its complex with adamantane-1-carboxylic acid showed that the hydroxy proton chemical shifts, temperature coefficients and vicinal coupling constants could be used to monitor the formation of intermolecular hydrogen bonds and hydration changes. The conformational change undertaken by altrose upon addition of adamantane-1-carboxylic acid allowed a more regular hydrogen bond network between the secondary hydroxyl groups in the CD, as evidenced by the downfield shift for O(3)H of the glucose-units. In the second part of the thesis, the interactions between di- and trimannosides, substructures of oligomannose-9, and mutants of the HIV inactivating protein cyanovirin-N (CV-N) were studied using saturation transfer difference NMR spectroscopy. In one mutant CV-NMutDB, the carbohydrate-binding site on domain B was suppressed while keeping the domain A intact. In the other mutant, CV-NMutDA, the specificity of domain A for trimannose was altered while domain B was kept intact. Both mutants recognised all di- and trimannosides containing the terminal Manα(1-2)Man epitope. The binding of the mutants were similar, with a slightly stronger affinity for the trisaccharide, Manα(1-2)Manα(1-2)ManαOMe in CV-NMutDB. Isothermal titration calorimetry data for CV-NMutDB showed a binding affinity of 3.4 +/-0.5 microM for the trisaccharide, which is close to the value derived for the nonamannoside (4.3 +/-0.5 microM). No binding parameters were extracted for CV-NMutDA due to the presence of two binding sites. This study confirm previous findings showing that not only the terminal disaccharide but also the linkage to the reducing end residue is important for binding to CV-N and thereby also for antiviral activity.