As part of our ongoing program aimed at understanding the structural significance of GlcNAcβAsn linkage conserved in all eukaryotic N-glycoproteins, the present study reports on the synthesis and X-ray crystal structures of N-(3-deoxy-3-acetamido-β-D-glycopyranosyl)acetamide (Glc 3NAcβNHAc) and the corresponding propionamide (Glc 3NAcβNHPr). Comparative analysis of these structures with those of the corresponding GlcNAc (C2 acetamido) compounds showed that the bifurcated anti-parallel pattern involving N-H-O and C-H-O hydrogen bonds, the hallmark feature of the N-glycoprotein models, GlcNAcβNHAc and GlcNAcβAsn, is absent in both the C3 acetamido analogs. The extended (anti) conformation of the amido aglycon moiety as defined by X2 seen in the case of C2 acetamido derivative, GlcNAcβNHPr, turns into gauche for the C3 acetamido analog (Glc3NAcβNHPr). This observation is consistent with the earlier work on the critical role of the C2-NHAc group of GlcNAcβAsn in controlling ?2 at the linkage region of N-glycoproteins. © The Author(s) 2011.

Manoharan Mathiselvam

Babu Varghese

Duraikkannu Loganathan

GLUCOSIDE

glycoprotein

article

chemical structure

chemistry

conformation

hydrogen bond

synthesis

X ray crystallography

Carbohydrate Conformation

Crystallography, X-Ray

Glucosides

glycoproteins

Hydrogen Bonding

Models, Molecular

IIT Madras is a public technical and research university located in Chennai, Tamil Nadu. Founded in 1959, it is recognised as an Institute of National Importance.

IIT Madras has been ranked as the top engineering institute in India for four years in a row by the National Institutional Ranking Framework of the MHRD

It currently offers undergraduate, postgraduate and research degrees across 16 disciplines in Engineering, Sciences, Humanities and Management. About 596 faculty belonging to science and engineering departments and centres of the Institute are engaged in teaching, research and industrial consultancy.

IIT Madras

Glycoconjugate Journal

The linkage region constituents, 2-deoxy-2-acetamido-β-D-glucopyranose (GlcNAc) and L-asparagine (Asn) are conserved in the N-glycoproteins of all eukaryotes. Elucidation of the structure and conformation of the linkage region of glycoproteins is important to understand the presentation and dynamics of the carbohydrate chain at the protein/cell surface. Earlier crystallographic studies using monosaccharide models and analogs of N-glycoprotein linkage region have shown that the N-glycosidic torsion, N, is more influenced by the structural variation in the sugar part than that of the aglycon moiety. To access the influence of distal sugar as well as interglycosidic linkage (α or β) on the N-glycosidic torsion angles, cellobiosyl and maltosyl alkanamides have been synthesized and structural features of seven of these analogs have been characterized by X-ray crystallography. Comparative analysis of the seven disaccharide analogs with the reported monosaccharide analogs showed that the N value of cellobiosyl analogs deviate ~9 with respect to GlcβNHAc. In the case of maltosyl analogs, deviation is more than 18. These deviations indicate that the N-glycosidic torsion is influenced by addition of distal sugar as well as with respect to inter glycosidic linkage (α or β); it is less influenced by changes occurring at the aglycon. The χ2 value of alkanamide derived from glucose, cellobiose and maltose exhibit a large range of variations (from 1.6 to -109.9). This large span of χ2 value suggests the greater degree of rotational freedom around C1′-C2′ bond which is restricted in GlcNAc alkanamides. The present finding explicitly proved the importance of molecular architecture in the N-glycoproteins linkage region to maintain the linearity, planarity and rigidity. These factors are necessary for N-glycan to serve role in inter- as well as intramolecular carbohydrate-protein interactions. © 2013 Springer Science+Business Media New York.

Effect of distal sugars and interglycosidic linkage on the N-glycoprotein linkage region conformation: Synthesis and X-Ray crystallographic investigation of β-1-N-alkanamide derivatives of cellobiose and maltose as disaccharide analogs of the conserved chitobiosylasparagine linkage

N-Linked glycosylation is not only present in eukaryotes but also occurs in archaea and bacteria and is mainly characterized by the β-glucosylamine linkage to the asparagine (GlcNAcβAsn). Earlier crystallographic studies aimed at understanding the structural significance of the linkage region constituents revealed that N-glycosidic torsion, φN is influenced considerably by variation in the glycan part as compared to the aglycon moiety. The φN value observed for XylβNHAc deviated maximum as compared to that of the model compound, GlcNAcβNHAc. The present work was undertaken to assess the influence of ribose on the N-glycosidic torsions and molecular assembly. Several ribopyranosyl alkanamides have been synthesized and crystal structures of three of them have been solved. A comprehensive crystal structure analysis of ribosyl alkanamides along with xylosyl and arabinosyl alkanamides showed the wide range of deviations in their φN values as compared to the negligible deviation shown by hexopyranosyl alkanamides. This study revealed the importance of C5-hydroxymethyl group and hydroxyl group configurations at C2, C3, and C4 stereocentres in controlling the N-glycosidic torsions. © 2013 Elsevier Ltd. All rights reserved.

Carbohydrate Research

Examination of the influence of C5-hydroxymethyl group and configurations of hydroxyl groups at C2, C3, and C4 stereocentres on the N-glycosidic torsion: Synthesis and X-ray crystallographic investigation of N-(d-ribopyranosyl) alkanamides as N-glycoprotein linkage region analogs

N-Glycoprotein linkage region constituents namely 2-deoxy-2-acetamido- β-d-glucopyranose (GlcNAc) and asparagine (Asn) are conserved among all eukaryotes. Earlier crystallographic studies on the linkage region conformation revealed that among all the models and analogs of the N-glycoprotein linkage region, XylβNHAc showed maximum deviation in the φN value as compared to the value reported for the model compound, GlcNAcβNHAc. In order to understand the effect of another pentopyranose, viz., arabinose, on the N-glycosidic torsion angles and molecular assembly, three arabinopyranosyl alkanamides were synthesized and their X-ray crystal structures elucidated. A comparative analysis of the N-glycosidic torsion, φN of the three analogs revealed the greater rotational freedom around the C1-N1 bond as compared to the GlcNAc derivatives. Molecular assembly of propionamido and chloroacetamido derivatives is characterized by the presence of anti-parallel bilayers of the molecules. This unique molecular assembly is hitherto unknown in all other models and analogs of N-glycoprotein linkage region. This study reveals that N-glycosidic torsions are influenced by the glycan as well as molecular packing. © 2013 Elsevier Ltd. All rights reserved.

Synthesis and X-ray crystallographic investigation of N-(α-d- arabinopyranosyl)alkanamides as N-glycoprotein linkage region analogs

N-Glycoprotein linkage region constituents, 2-acetamido-2-deoxy-β-D- glucopyranose (GlcNAc) and asparagine (Asn) are conserved among all the eukaryotes. To gain a better understanding for nature's choice of GlcNAcβAsn as linkage region constituents and inter- and intramolecular carbohydrate-protein interactions, a detailed systemic structural study of the linkage region conformation is essential. Earlier crystallographic studies of several N-(β-glycopyranosyl)alkanamides showed that N-glycosidic torsion, φN, is influenced to a larger extent by structural variation in the sugar part than that of the aglycon moiety. To explore the effect of the bioisosteric replacement of a carboxamide group by a sulfonamide moiety on the N-glycosidic torsions as well as on molecular assembly, several glycosyl methanesulfonamides and glycosyl chloromethanesulfonamides were synthesized as analogues of the N-glycoprotein linkage region, and crystal structures of seven of these compounds have been solved. A comparative analysis of this series of crystal structures as well as with those of the corresponding alkanamido derivatives revealed that N-glycosidic torsion, φN, does not alter significantly. Methanesulfonamido and chloromethanesulfonamido derivatives of GlcNAc display a different aglycon conformation compared to other sulfonamido analogues. This may be due to the cumulative effect of the direct hydrogen bonding between N1 and O1′ and C-H×××O interactions of the aglycon chain, revealing the uniqueness of the GlcNAc as the linkage sugar. Unique molecular assembly motif of GlcNAc: The different aglycon conformations of methanesulfonamido and chloromethanesulfonamido derivatives of GlcNAc as compared to other sulfonamido analogues is a unique feature of their molecular assembly. This could be due to the cumulative effect of the direct hydrogen bonding between N1 and O1′ and C-H×××O interactions of the aglycon chain. Copyright © 2013 WILEY-VCH Verlag GmbH &amp; Co. KGaA, Weinheim.

Chemistry - A European Journal

Exploring the effect of bioisosteric replacement of carboxamide by a sulfonamide moiety on N-glycosidic torsions and molecular assembly: Synthesis and x-ray crystallographic investigation of n-(β- D -glycosyl)sulfonamides as n-glycoprotein linkage region analogues

The linkage region, GlcNAcβAsn, is conserved in all eukaryotic N-glycoproteins. As a logical extension of a research endeavor aimed at understanding the structural significance of GlcNAc and Asn as the linkage region constituents, the newer analogs GlcNAcβNHBu and (GlcNAcβ(1-4) GlcNAc)alkanamides have been synthesized to assess the influence of aglycon as well as additional GlcNAc on the linkage region. X-ray crystallographic analysis of the GlcNAcβNHBu and (GlcNAcβ(1-4)GlcNAc)βNHBu is described. Comparative analysis of these structures with those of reported models and analogs shows that the deviation in N-glycosidic torsion, φN among the GlcNAc alkanamides is negligible (&lt;2) whereas (GlcNAcβ(1-4) GlcNAc)βNHBu deviates by ∼15 as compared to GlcNAcβNHBu. Under the influence of the molecular packing, the conformation around the C1′-C2′ bond deviates from anti to gauche in (GlcNAcβ(1-4) GlcNAcβNHBu. Interestingly, C2-acetamido group in (GlcNAcβ(1-4)GlcNAc) NHBu orients differently as compared to GlcNAc alkanamides and this orientation was found to be almost similar to β-N,N′-diacetylchitobiose trihydrate. The bifurcated anti-parallel pattern involving N-HO and C-HO hydrogen bonds, a hallmark feature of the N-glycoprotein models, GlcNAcβNHAc and GlcNAcβAsn, is absent in both the title alkanamides. This is the first report on the crystal structure analysis of chitobiosyl alkanamide as analog of the N-glycoprotein linkage region, (GlcNAcβ(1-4) GlcNAc)βAsn. © 2013 Published by Elsevier Ltd.

Synthesis and X-ray crystallographic investigation of N-(β-d-glycosyl) butanamides derived from GlcNAc and chitobiose as analogs of the conserved chitobiosylasparagine linkage of N-glycoproteins

The biological addition of oligosaccharide moieties to asparagine residues of N-glycoproteins influences the properties and bioactivities of these macromolecules. The low-temperature neutron crystal structures of three N-glycoprotein linkage models and analogues provide accurate characterization of the three-dimensional structure of the conserved GlcNAc - Asn linkage. These first crystal structures of N-acetylated carbohydrates obtained by neutron diffraction provide high-resolution geometrical parameters that can be used for force-field parametrization and subsequent molecular dynamics simulation of N-glycoproteins. The correct localization of hydrogen atoms demonstrates the occurrence of trifurcated hydrogen bonds and hydrophobic contacts. © 2011 American Chemical Society.

Journal of the American Chemical Society

Low-temperature neutron diffraction structures of N -glycoprotein linkage models and analogues: Structure refinement and trifurcated hydrogen bonds

Elucidation of the intra- and intermolecular carbohydrate-protein interactions would greatly contribute toward obtaining a better understanding of the structure-function correlations of the protein-linked glycans. The weak interactions involving C-H...O have recently been attracting immense attention in the domain of biomolecular recognition. However, there has been no report so far on the occurrence of C-H...O hydrogen bonds in the crystal structures of models and analogs of N-glycoproteins. We present herein an analysis of C-H...O interactions in the crystal structures of all N-glycoprotein linkage region models and analogs. The study reveals a cooperative network of bifurcated hydrogen bonds consisting of N-H...O and C-H...O interactions seen uniquely for the models. The cooperative network consists of two antiparallel chains of bifurcated hydrogen bonds, one involving N1-H, C2′-H and O1′ of the aglycon moiety and the other involving N2-H, C1-H and O1″ of the sugar. Such bifurcated hydrogen bonds between the core glycan and protein are likely to play an important role in the folding and stabilization of proteins. © 2006 Oxford University Press.

Fulltext

Glycobiology

Observation of a unique pattern of bifurcated hydrogen bonds in the crystal structures of the N-glycoprotein linkage region models

The linkage region constituents, namely, 2-acetamido-2-deoxy-β-D- glucopyranose and asparagine are conserved in the N-glycoproteins of all the eukaryotes. The present work is aimed at understanding the reasons for the occurrence of GlcNAc and Asn as the linkage region constituents. A total of six sugar amides have been designed as models and analogs of the linkage region and their crystal structures have been solved. This is the first report on the X-ray crystallographic investigation of the effect of systematic changes in the linkage sugar as well as its aglycon moiety on the N-glycosidic torsion, ψN (O5-C1-N1-C1′). This also forms the first report on the crystal structure of a model of L-RhaβAsn, a variant linkage found in the surface layer glycoprotein of Bacillus stearothermophillus. Among the models and analogs examined, the acetamido derivatives of Man and Xyl, the linkage sugars of O-glycoproteins, show a ψN value of -114. 5° and -121.2°, respectively, deviating maximum from the value of -89.8° reported for the model compound GlcNAcβNHAc. The L-Rha and Gal derivatives also show noticeable deviations. The ψN values, -89.5° and -91.0°, of the propionamide derivatives of Glc and GlcNAc (analogs of GlcβGln and GlcNAcβGln, respectively) agree well with those (-93.8° and -89.8°) reported for their corresponding acetamide derivatives suggesting Gln could serve as well as Asn as the linkage region amino acid. However, the rotational freedom about the additional C-C bond would lead to altered rigidity of the linkage region. An analysis of packing reveals that the molecular assembly of these compounds is driven by different infinite and finite chains of hydrogen bonds. The double pillaring of hydrogen bonds involving the amide groups at C1 and C2 is seen as a unique packing feature characteristic of β-1-N-acyl derivatives of GlcNAc. Based on the findings of the present study, it is speculated that the linkage region constituents of the eukaryotic N-glycoproteins appear to fulfill three essential structural requirements: rigidity, planarity, and linearity and these are met by the trisaccharide core and Asn at the linkage region. © 2003 Elsevier Inc. All rights reserved.

Biochemical and Biophysical Research Communications

On the structural significance of the linkage region constituents of N-glycoproteins: An X-ray crystallographic investigation using models and analogs

X-Ray diffraction analysis of a hydrated N-glycoprotein linkage region model, N1-(2-acetamido-2-deoxy-β-D-glucopyranosyl)acetamido- 2-deoxy-β-D-glucopyranosyl)acetamide (1) and its N1-benzamido analogue 2 reveals very interesting molecular architecture involving seven hydrogen bonds in both compounds and further stabilization of packing in 2 due to π-π stacking. The crystal structure has a two dimensional network of hydrogen bonds formed by infinite chains running along the b-axis and homodromic cycles, intersecting at the water oxygens.

Crystal structure of a hydrated N-glycoprotein linkage region model and its analogue: Hydrogen bonding and π-π stacking driven molecular assembly

Biochemistry

The Expanding Horizons of Asparagine-Linked Glycosylation

Synthesis and X-ray crystallographic investigation of N-(3-deoxy-3- acetamido-β-D-glycopyranosyl)alkanamides as analogs of N-glycoprotein linkage region

Journal	Glycoconjugate Journal
ISSN	02820080
Open Access	No