New NMR Tools for Characterizing the Dynamic Conformations and Interactions of Oligosaccharides

Abstract

Oligosaccharides play pivotal roles in various molecular recognition events on cell surfaces and in intracellular environments. Although information about the three-dimensional structure of oligosaccharides is crucial for understanding the mechanisms underlying their biological functions and for designing drugs targeting carbohydrate recognition systems, their inherent flexibility hampers detailed conformational analysis. NMR spectroscopy has immense potential for providing atomic details of oligosaccharide structures in solution as well as in complexes with other biomolecules. However, the traditional NMR approach based on local conformational information provided by the nuclear Overhauser effect (NOE) is often precluded by insufficient information about long interatomic distances and hydrogen bonds involving hydroxy groups. To address these issues, new NMR techniques have recently been developed, exploiting the effects of introduced paramagnetic probes and stable isotopes to target oligosaccharides. Lanthanoid tagging and the spin labeling of oligosaccharides induce paramagnetic effects such as pseudocontact shift and paramagnetic relaxation enhancement, offering NOEindependent sources of long-distance information. Deuteriuminduced isotope shifts of neighboring 13 C resonances provide an experimental tool for the identification of hydrogen bonds involving oligosaccharide hydroxy groups. The uniform and position-selective 13 C enrichment of oligosaccharides can be achieved by metabolic labeling using genetically engineered yeast cells. Furthermore, small ganglioside-embedding bicelles have been used for detailed NMR analyses of intermolecular interactions involving glycolipids in membrane environments. These NMR approaches, in conjunction with computational simulation, have opened up new vistas for the analysis of oligosaccharide conformations and interactions at atomic level. Ç IntroductionOligosaccharides, which are formed by the combination of approximately ten types of monosaccharides through various glycosidic linkages, are a major class of essential molecular components in biological systems. By modifying proteins and lipids, oligosaccharides serve as mediators in physiological and pathological molecular recognition events; thus, they are involved in a wide range of cellular processes including proteinfate determination, intercellular communication, and viral infections.1 The detailed characterization of the 3D structures of oligosaccharides in both free and bound states is crucial for understanding their underlying functions and mechanisms and for providing insights into recognition mechanisms that are useful in the design of carbohydrate-based drugs. 2,3Compared to protein structural biology, 3D-structural study of oligosaccharides is still immature, primarily because of high inherent flexibility. The many degrees of freedom in internal motion exhibited by oligosaccharides endow them with conformational adaptability in interacting with various target biomolecules. 2,4 This makes the structural a...

Recent Advances in NMR Studies of Carbohydrates

“…Few years back, several comprehensive reviews about NMR techniques were published. In 2013, Kato and coworkers [1] presented an excellent review about new NMR tools for characterizing the dynamic conformation and interactions of oligosaccharides. A comprehensive overview of the advances in glycosaminoglycanomics such as chondroitin sulfate, dermatan sulfate, heparin, heparan sulfate, and hyaluronan by 15 N NMR spectroscopy has been written by Pomin [2].…”

“…This method allows to follow the reaction progress and determines the purity of isolated compounds. 1 H NMR spectra of carbohydrates can be distinguished to some characteristic including those of anomeric protons ($δ 4.4-5.5), acetyl ($δ 2.0-2.1), and methyl ($δ 1.2) groups, and other protons that are influenced by specific functionality, including amino groups, phosphorylation, sulfation, glycosylation, and acetylation, or the lack of functionality as in deoxysugars. One of the most important NMR experiments for carbohydrate study is WATERGATE (WATER suppression by GrAdient Tailored Excitation).…”

“…The 1 H, 13 C, 31 P NMR spectra have been recorded at in D 2 O at 70°C. Chemical shifts were assigned performed using 1D 1 H and 13 C experiments together with a number of 2D 1 H, 1 Fig. 7 Conformation distribution due to protonation.…”

Paramagnetic NMR probes for characterization of the dynamic conformations and interactions of oligosaccharides

“…The polar coordinates of the nucleus, r, θ, and φ, are defined with respect to the paramagnetic center and the principal axis of the Δχ tensor. Adapted from Zhang et al 2013 [12] with permission from The Chemical Society of Japan Paramagnetic probes of biomolecular NMR Appropriate paramagnetic probes, i.e., paramagnetic metal ions or spin labels, are required to observe paramagnetic effects. Paramagnetic NMR spectroscopy has traditionally been developed for the characterization of metalloproteins because they possess endogenous paramagnetic probes and have intrinsic metal ion-binding sites that can be used to incorporate paramagnetic ions [32,33].…”

“…Paramagnetism-assisted NMR approaches have recently been rekindled in the fields of NMR structural biology, including carbohydrate structural studies [12][13][14][15]. This revival has occurred in the context of the development of molecular biology-based sample preparation, in conjunction with the development of chemically designed paramagnetic probes as intelligence sources [16][17][18][19][20][21][22][23].…”