Solution and Solid-State NMR Studies to Advance the Application of MA’AT Analysis for Structural and Conformational Analysis of Saccharides

Carbohydrates have always been known to play key roles in energy metabolism and biological structure, but in recent decades they have been shown to play major roles in drug discovery, cell signaling, and biomolecular recognition. These important biological functions of saccharides are largely attributed to molecular flexibility and a diverse set of structural components. Experimental and theoretical techniques for modeling saccharide conformation and structure need continued development to aid in the understanding of biological functions of glycans.

The Serianni laboratory has been developing MA’AT Analysis to model saccharide conformation in solution. The method relies on redundant NMR J-values and density functional theory (DFT) calculations to probability distributions of molecular torsion angles in aqueous environments and circular standard deviations (CSDs) to evaluate librational motions about these torsion angles. Chapter 2 describes the use of MA’AT analysis to model N-acetyl side-chains of common saccharides. Chapter 5 describes the use of MA’AT analysis on the Glc₃Man tetrasaccharide and its representative disaccharides in isolation, which are fragments of the parent N-glycan Glc₃Man₉GlcNAc₂. The results from Chapter 5 are to determine the context effects that the Glc₃Man tetrasaccharide has on its individual O-glycosidic linkages, to determine the ability of MA’AT Analysis to detect the presence of two major conformers, and increase the robustness of the MA’AT method. Chapter 6 describes the synthesis of a Man₃ trisaccharide and Man₃GlcNAc₂ pentasaccharide for future plans of conducting MA’AT Analysis of large complex-type N-glycans.

Chapters 3 and 4 deal with determining whether direct spin-spin couplings can be used as experimental variables in MA’AT Analysis. Chapter 3 describes the modeling of conformationally-constrained saccharides in aqueous solution to observe the difference between experimental J-couplings and those calculated via density functional theory (DFT). Chapter 4 describes the modeling of monosaccharides in the solid-state to observe the difference between experimental solid-state J-couplings and those calculated from DFT. Both chapters are connected in reaffirming each other’s findings that while DFT quantitatively predicts J-values within reasonable error, DFT functionals and basis sets need further development to properly capitulate ¹J values for use in MA’AT analysis.