(W1130-04-23) Synthesis, Conformational Analysis, and Binding Studies of 6α-mannobiose Glycosides

Purpose: The interactions of sugars with protein receptors mediates a plethora of biological functions including immune response, fertilization, metastasis, infection, among many others. For the protein-carbohydrate interactions to be successful it is necessary for the carbohydrate molecule to be in a specific conformation that is imposed by its protein receptor. Thus, the study of the conformational preferences of saccharides is a fundamental factor for understanding protein-carbohydrate interactions. The free rotation around the inter-glycosidic bond and the hydroxymethyl group (C5-C6) are the main factors determining the conformational flexibility of saccharides, especially those containing (1-6) connections. Various factors are known to determine the rotation around C5-C6, among them the type of substitution on hydroxyl C6, the configuration of the anomeric carbon, the media polarity, and the configuration of the stereocenter C4. Moreover, previous studies on monosaccharide glycosides showed that the conformational preferences around C5-C6 are also determined by the chemical nature of the aglycone and the stereochemical configuration of the anomeric center. From the study it was observed that changes on the aglycone’s steric properties lead to different rotamer contributions around C5-C6. Thus, increasing the aglycone’s size from a methyl to a tert-butyl group lead to increasing gt contributions at the cost of gg.¹ To date, no such studies have been performed on disaccharides or higher saccharides. In the present communication, we disclose our preliminary results on the NMR-based conformational analysis of a series of alkyl 6α-mannobiosides in solution. These compounds were designed to explore in larger saccharides the structure-conformation relationships found in monosaccharides.

Methods: To better understand the effect of aglycone substituent over the conformational properties around C5-C6 bond in higher saccharides, we designed and synthesized a series α and β mannosides and 6α-mannobiosides (aManOR , aMan(1→6) aManOR, bManOR, aMan(1→6) bManOR) of aliphatic alcohols. We will study the conformational flexibility around the interglycosidic bond by calculating the rotamer distribution around C5-C6 using the ³J_H5,H6R and ³J_H5,H6S coupling constants and the set of equations developed by Serianni.
2.8 Pgg + 2.2 Pgt + 11.1 Ptg = ³J_H5H6S
0.9 Pgg + 10.8 Pgt + 4.2 Ptg = ³J_H5H6R
Pgg + Pgt + Ptg = 1
From the conformational study, we expect to establish structure-conformation relationships that will help for a better understanding of carbohydrate recognition mechanisms and could assist in the design of conformationally tailored ligand candidates in the future. Next, we will focus on studying the relationship between flexibility and binding affinity using the mannose-binding lectin concanavalin A.

Results: From the NMR analysis (Table 1) of a first series of protected aMan(1→6) aManOR glycosides, we found no significant changes on the interglycosidic conformation among glycosides as a function of the aglycon’s structure. Most compounds exhibit a 50% contribution of the gg and gt rotamers to the rotational equilibrium about C5-C6. However, the glycoside substituted with the bulkier aglycone, tert-butyl derivative 1f, exhibits larger gt contribution in a conformational equilibrium with equal gg and gt distribution in the other glycosides (Table 2). The result obtained contrasts from the conformational properties of glucose monosaccharides and our previous studies in rutinosides.^{1 2} However, it is in alignment with the results obtained from previous studies performed on mannose monosaccharide molecules where no change in conformation for a mannosides is observed.³ However, given the background knowledge on β-mannosides, where alkyl aglycones modulate the C5-C6 conformation,⁴ we will focus on synthesizing and analyzing β-glycosides of 6α-mannobiose. We expect to find in these substrates the structure-conformation relationship seen on β-glucosides and β-mannosides and correlate the conformational flexibility with the binding affinity towards a biological receptor such as the mannose-binding protein concanavalin A.

Conclusion: In contrast to glucosides and mannosides, the C5-C6 flexibility of α-glycosides of Man(1,6)Man disaccharides is not significantly influenced by the nature of the aglycone. With exception of tert-butyl derivative, all 6α-mannobiose glycosides exhibit the same rotamer distribution around the interglycosidic linkage. To further explore the effect of the anomeric configuration about the C5-C6 flexibility, we will continue this study focusing on the conformational properties of the respective β-glycosides.

References: (1) Sanhueza, C. A.; Dorta, R. L.; Vázquez, J. T. Involvement of the S-Aglycon in the Conformational Preferences of Thioglucosides. Tetrahedron: Asymmetry 2008, 19 (2), 258–264. https://doi.org/10.1016/j.tetasy.2008.01.003.
(2) Mehta, D. A. Agal Antigen and Rutinose Glycosides as Model Compounds for the Design of Classical and Conformational Glycomimetics. 133.
(3) Nóbrega, C.; Vázquez, J. T. Conformational Study of the Hydroxymethyl Group in α-d-Mannose Derivatives. Tetrahedron: Asymmetry 2003, 14 (18), 2793–2801. https://doi.org/10.1016/S0957-4166(03)00623-2.
(4) Mayato, C.; Dorta, R.; Vázquez, J. Experimental Evidence on the Hydroxymethyl Group Conformation in Alkyl β-d-Mannopyranosides. Tetrahedron: Asymmetry 2004, 15 (15), 2385–2397. https://doi.org/10.1016/j.tetasy.2004.06.019.

Acknowledgements: We thank St. John's University College of Pharmacy and health Sciences for funding our research