Unexpected structure of a C. difficile toxin A ligand necessitates an annotation correction in a popular screening library
Department of Chemistry, Alberta Ingenuity Centre for Carbohydrate Science, University of Calgary, Calgary AB T2N1N4, Canada. Chemical Communications
(Impact Factor: 6.83).
12/2011; 47(45):12397-9. DOI: 10.1039/c1cc15344g
The structure of the pentasaccharide S259-1 in the Consortium for Functional Glycomics was investigated using a variety of techniques. Surprisingly, the structure differs from the structure assumed from the previously established specificity of the human fucosyltransferase FUT-III used in the last step of chemoenzymatic synthesis. When presented with a tetrasaccharide substrate containing both type I and type II disaccharide moieties, the enzyme generates a pentasaccharide in which the type II moiety is preferentially fucosylated. The unexpected product generated by FUT-III in this case highlights the importance of performing detailed structural analysis on products generated by enzymes.
Available from: Chang-Chun Ling
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ABSTRACT: We report the synthesis of a tetrasaccharide and two pentasaccharide fragments of the LeaLex tumor associated carbohydrate antigen alpha-L-Fuc-(1->4)-[beta-D-Gal-(1->3)]-beta-D-GlcNAc-(1->3)-beta-D-Gal-(1->4)-[alpha-L-Fuc-(1->3)]-beta-D-GlcNAc-(1->OR). The choice of protecting groups permitted a one-step global deprotection (Na/NH3(l)). The protected chlorohexyl glycoside pentasaccharide was the precursor to the hexyl glycoside, to be used as a soluble inhibitor, and the aminohexyl glycoside analogue, to be conjugated to proteins for surface immobilization and immunization experiments. We observed that a linear tetrasaccharide that contained two N-acetylglucosamine residues and a free OH group gave two distinct sets of 1H NMR signals when the data was acquired in deuterated chloroform. Data acquisition at variable concentrations and variable temperatures suggest that the second set of NMR signals results from aggregation of the tetrasaccharide driven by the formation of intermolecular H-bonds involving the NHAc. While the formation of intra- and intermolecular H-bonds involving N-acetylgucosamine residues has been reported in non-H-bonding solvents, this is, to our knowledge, the first time that these lead to the appearance of two distinct sets of signals in the NMR spectra. This aggregation, may explain the lack of reactivity observed when attempting to glycosylate such acceptor using non H-bonding solvents such as dichloromethane.
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ABSTRACT: We report the efficient synthesis of three analogues of the tumor associated carbohydrate antigen LeaLex. This hexasaccharide was prepared as a soluble inhibitor hexyl glycoside, a 6-aminohexyl glycoside for conjugation to proteins, and as 6-thiohexyl glycoside for immobilization to a gold surface. These three analogues were obtained from a common hexasaccharide intermediate and isolated pure following efficient deprotection reactions that involved metal dissolving conditions. While all other intermediates and analogues gave the expected molecular ions in ESI HRMS, the 6-thiohexyl glycoside final compound gave a complex spectrum in which no signal matched the molecular ion. Using ESI FAIMS HRMS, we were able to prevent ion dissociation reactions and obtained high quality spectral data. The ions detected could be characterized unambiguously from their accurate masses and gave insight into the behavior of the thiohexyl analogue in the gas phase. These results indicate that the 6-thiohexyl glycoside lost water and led to the formation of "hyper-metalated" species which we propose are cyclic.
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