Related Disaccharides Research Articles

A Simple Biomimetic Receptor Selectively Recognizing the GlcNAc2 Disaccharide in Water.

GlcNAc2 is the core disaccharide fragment present in N‐glycans exposed on the surface of enveloped viruses of high health concern, such as coronaviruses. Because N‐glycans are directly involved in the docking of viruses to host cells, recognition of GlcNAc2 by a biomimetic receptor may be a convenient alternative to the use of lectins to interfere with viral entry and infection. Herein, we describe a simple biomimetic receptor recognizing the methyl‐β‐glycoside of GlcNAc2 in water with an unprecedented affinity of 160 μM, exceeding that of more structurally complex receptors reported in the literature. The tweezers‐shaped acyclic structure exhibits marked selectivity among structurally related disaccharides, and complete discrimination between mono‐ and disaccharides. Molecular modelling calculations supported by NOE data provided a three‐dimensional description of the binding mode, shedding light on the origin of the affinities and selectivities exhibited by the receptor.

A Simple Biomimetic Receptor Selectively Recognizing the GlcNAc2 Disaccharide in Water

AbstractGlcNAc2 is the core disaccharide fragment present in N‐glycans exposed on the surface of enveloped viruses of high health concern, such as coronaviruses. Because N‐glycans are directly involved in the docking of viruses to host cells, recognition of GlcNAc2 by a biomimetic receptor may be a convenient alternative to the use of lectins to interfere with viral entry and infection. Herein, we describe a simple biomimetic receptor recognizing the methyl‐β‐glycoside of GlcNAc2 in water with an unprecedented affinity of 160 μM, exceeding that of more structurally complex receptors reported in the literature. The tweezers‐shaped acyclic structure exhibits marked selectivity among structurally related disaccharides, and complete discrimination between mono‐ and disaccharides. Molecular modelling calculations supported by NOE data provided a three‐dimensional description of the binding mode, shedding light on the origin of the affinities and selectivities exhibited by the receptor.

Generation of β-Linked GlcNac and GalNAc Terminating Disaccharide Library through Glycosylation on Unprotected Monosaccharides

Background: The GlcNAc related sulfated disaccharides were reported to show inhibitory activity on breast tumor cells (MDA-MB-231). However, synthesis of sulfated disaccharides is laborious and time consuming, and there is a need to find more efficient methods to generate the sulfated disaccharide libraries in acceptable ratios and in respectable yields. Random glycosylation followed by sulfation may be a good method to synthesize sulfated disaccharide library. However, there is a dearth of simple methods to generate the GalNAc and GlcNAc related disaccharides with acceptable ratio by random glycosylation. Results: Following a generic procedure, random glycosylation of unprotected monosaccharides and followed by deprotection, acetylation, deacetylation, and purification, yielded the GalNAc-GlcNAc disaccharide library containing three β-linked disaccharides with a 1:1:1 ratio and the GlcNAc-Gal disaccharide library containing four β-linked disaccharides with 1:1:2:2 ratio. These results were confirmed by comparison of their proton NMR spectra with that of standard disaccharides. Conclusion: Random glycosylation was employed to generate GalNAc and GlcNAc related disaccharide libraries and was demonstrated as the most efficient method thus far for the synthesis of GalNAc and GlcNAc related disaccharide libraries with excellent ratios, which further can be used as the starting materials for random sulfation to synthesize the sulfated disaccharide libraries. Keywords: Oligosaccharide library, synthesis, combinatorial chemistry, GalNAc-GlcNAc disaccharides, GlcNAc-Gal disaccharides, glycoprotein.

Generation of α-Linked Oligosaccharide Libraries by Random Glycosylation on Unprotected Acceptors

Background: The GlcNAc related disaccharides were reported to show inhibitory activity on breast tumor cells (MDAMB- 231). However, synthesis of disaccharides is lengthy, laborious and time consuming, and it is necessary to find more efficient methods to generate the disaccharide libraries. Random glycosylation has been an established practice to generate β-linked disaccharides library. Synthesis of α-linked disaccharide libraries by random glycosylation hasn’t been explored yet. We present our results on the synthesis of α-linked GlcNAc related disaccharide libraries by random glycosylation in this article. Results: Employing random glycosylation as a key step on an unprotected GlcNAc monosaccharide, and after deprotection and careful purification, few α-linked GlcNAc related disaccharide libraries were obtained in acceptable ratios, and the results were confirmed by mass and NMR spectral data. Conclusion: Random glycosylation was demonstrated as an efficient method for the synthesis of α-linked GlcNAc related oligosaccharide libraries.

Synthesis of 2-deoxy ribose related disaccharide nucleoside and its phosphoramidite

Synthesis of impurity reference compound of anti-tumor drug ISIS 183750 was achieved. In this process, a general method for synthesis of 2-deoxy ribosyl disaccharide nucleosides was established for the first time.

Identification of structurally closely related monosaccharide and disaccharide isomers by PMP labeling in conjunction with IM-MS/MS.

It remains particularly difficult for gaining unambiguous information on anomer, linkage, and position isomers of oligosaccharides using conventional mass spectrometry (MS) methods. In our laboratory, an ion mobility (IM) shift strategy was employed to improve confidence in the identification of structurally closely related disaccharide and monosaccharide isomers using IMMS. Higher separation between structural isomers was achieved using 1-phenyl-3-methyl-5-pyrazolone (PMP) derivatization in comparison with phenylhydrazine (PHN) derivatization. Furthermore, the combination of pre-IM fragmentation of PMP derivatives provided sufficient resolution to separate the isomers not resolved in the IMMS. To chart the structural variation observed in IMMS, the collision cross sections (CCSs) for the corresponding ions were measured. We analyzed nine disaccharide and three monosaccharide isomers that differ in composition, linkages, or configuration. Our data show that coexisting carbohydrate isomers can be identified by the PMP labeling technique in conjunction with ion-mobility separation and tandem mass spectrometry. The practical application of this rapid and effective method that requires only small amounts of sample is demonstrated by the successful analysis of water-soluble ginseng extract. This demonstrated the potential of this method to measure a variety of heterogeneous sample mixtures, which may have an important impact on the field of glycomics.

Crystal packing in three related disaccharides: precursors to heparan sulfate oligosaccharides.

The three title compounds form part of a set of important precursor dissacharides which lead to novel therapeutics, in particular for Alzheimer's disease. All three crystallize as poorly diffracting crystals with one independent mol-ecule in the asymmetric unit. Two of them are isostructural: 4-meth-oxy-phenyl 4-O-[6-O-acetyl-2-azido-3-O-benzyl-2-de-oxy-4-O-(9-fluor-en-yl-methyl-oxycarbon-yl)-α-d-gluco-pyranos-yl]-2-O-benzoyl-3-O-benzyl-6-O-chloro-acetyl-α-l-ido-pyran-oside, C59H56ClN3O16, (I), the ido-relative of a reported gluco-disaccharide [Gainsford et al., 2013 ▶). Acta Cryst. C69, 679-682] and 4-meth-oxy-phenyl 4-O-[6-O-acetyl-2-azido-3-O-benzyl-2-de-oxy-4-O-(9-fluorenyl-methyl-oxycarbon-yl)-α-d-gluco-pyranos-yl]-2-O-benzoyl-3-O-benzyl-6-O-meth-oxy-acetyl-α-l-ido-pyran-oside, C60H59N3O17, (II). Both exhibit similar conformational disorder of pendant groups. The third compound 4-meth-oxy-phenyl 4-O-[6-O-acetyl-2-azido-3,4-di-O-benzyl-2-de-oxy-α-d-gluco-pyranos-yl]-2-O-benzoyl-3-O-benzyl-6-O-meth-oxy-oacetyl-β-d-gluco-pyran-oside, C52H55N3O15, (III), illustrates that a slightly larger set of weak inter-molecular inter-actions can result in a less disordered mol-ecular arrangement. The mol-ecules are bound by weak C-H⋯O(ether) hydrogen bonds in (I) and (II), augmented by C-H⋯π inter-actions in (III). The absolute configurations were determined, although at varying levels of significance from the limited observed data.

Sequence-constructive SELEX: A new strategy for screening DNA aptamer binding to Globo H

We proposed to use a novel stepwise sequence-constructive SELEX method to develop DNA aptamers that can recognize Globo H which is a tumor-associated carbohydrate antigen. A combinatorial synthetic library that consisted of DNA molecules with randomized regions of 15-bases was used as the starting library for the first SELEX procedure. The input DNA library for the second round of SELEX consisted of the extension of the 5′ and 3′-ends with 7-bases that were randomized from four selected aptamers. The third round of SELEX was performed following the same procedures as described for the second round of SELEX. The experimental results indicate that the binding affinity of DNA aptamers to Globo H was enhanced when using the sequence-constructive SELEX approach. The selectivity of the DNA aptamers for related disaccharides, mannose derivatives, and Globo H analogs demonstrated the ability of the DNA aptamers to discriminate the presence of various glycans with different structures.

Methyl 4-O-β-D-xylopyranosyl β-D-mannopyranoside, a core disaccharide of an antifreeze glycolipid

Methyl β-D-xylopyranosyl-(1→4)-β-D-mannopyranoside, C12H22O10, crystallized as colorless block-like needles from methanol-water solvent. Comparisons to the internal linkage conformations in the two crystallographic forms of the structurally related disaccharide methyl β-D-mannopyranosyl-(1→4)-β-D-xylopyranoside are discussed. Intramolecular inter-residue hydrogen bonding is observed between one mannopyranosyl hydroxy O atom and the ring O atom of the xylopyranosyl residue. Intermolecular hydrogen bonding yields a bilayered two-dimensional sheet of molecules that are located parallel to the bc plane.

Methyl 4-O-β-D-mannopyranosyl β-D-xylopyranoside

Methyl β-D-mannopyranosyl-(1→4)-β-D-xylopyranoside, C(12)H(22)O(10), (I), crystallizes as colorless needles from water, with two crystallographically independent molecules, (IA) and (IB), comprising the asymmetric unit. The internal glycosidic linkage conformation in molecule (IA) is characterized by a ϕ' torsion angle (O5'(Man)-C1'(Man)-O1'(Man)-C4(Xyl); Man is mannose and Xyl is xylose) of -88.38 (17)° and a ψ' torsion angle (C1'(Man)-O1'(Man)-C4(Xyl)-C5(Xyl)) of -149.22 (15)°, whereas the corresponding torsion angles in molecule (IB) are -89.82 (17) and -159.98 (14)°, respectively. Ring atom numbering conforms to the convention in which C1 denotes the anomeric C atom, and C5 and C6 denote the hydroxymethyl (-CH(2)OH) C atom in the β-Xylp and β-Manp residues, respectively. By comparison, the internal glycosidic linkage in the major disorder component of the structurally related disaccharide, methyl β-D-galactopyranosyl-(1→4)-β-D-xylopyranoside), (II) [Zhang, Oliver & Serriani (2012). Acta Cryst. C68, o7-o11], is characterized by ϕ' = -85.7 (6)° and ψ' = -141.6 (8)°. Inter-residue hydrogen bonding is observed between atoms O3(Xyl) and O5'(Man) in both (IA) and (IB) [O3(Xyl)...O5'(Man) internuclear distances = 2.7268 (16) and 2.6920 (17) Å, respectively], analogous to the inter-residue hydrogen bond detected between atoms O3(Xyl) and O5'(Gal) in (II). Exocyclic hydroxymethyl group conformation in the β-Manp residue of (IA) is gauche-gauche, whereas that in the β-Manp residue of (IB) is gauche-trans.

Disorder and conformational analysis of methyl β-D-galactopyranosyl-(1→4)-β-D-xylopyranoside

Methyl β-D-galactopyranosyl-(1→4)-β-D-xylopyranoside, C(12)H(22)O(10), (II), crystallizes as colorless needles from water with positional disorder in the xylopyranosyl (Xyl) ring and no water molecules in the unit cell. The internal glycosidic linkage conformation in (II) is characterized by a φ' torsion angle (C2'(Gal)-C1'(Gal)-O1'(Gal)-C4(Xyl)) of 156.4 (5)° and a ψ' torsion angle (C1'(Gal)-O1'(Gal)-C4(Xyl)-C3(Xyl)) of 94.0 (11)°, where the ring atom numbering conforms to the convention in which C1 denotes the anomeric C atom, and C5 and C6 denote the hydroxymethyl (-CH(2)OH) C atoms in the β-Xyl and β-Gal residues, respectively. By comparison, the internal linkage conformation in the crystal structure of the structurally related disaccharide, methyl β-lactoside [methyl β-D-galactopyranosyl-(1→4)-β-D-glucopyranoside], (III) [Stenutz, Shang & Serianni (1999). Acta Cryst. C55, 1719-1721], is characterized by φ' = 153.8 (2)° and ψ' = 78.4 (2)°. A comparison of β-(1→4)-linked disaccharides shows considerable variability in both φ' and ψ', with the range in the latter (~38°) greater than that in the former (~28°). Inter-residue hydrogen bonding is observed between atoms O3(Xyl) and O5'(Gal) in the crystal structure of (II), analogous to the inter-residue hydrogen bond detected between atoms O3(Glc) and O5'(Gal) in (III). The exocyclic hydroxymethyl conformations in the Gal residues of (II) and (III) are identical (gauche-trans conformer).

Methyl β-D-galactopyranosyl-(1→4)-β-D-allopyranoside tetrahydrate

The title compound, C13H24O11·4H2O, (I), crystallized from water, has an inter­nal glycosidic linkage conformation having φ′ (O5Gal—C1Gal—O1Gal—C4All) = −96.40 (12)° and ψ′ (C1Gal—O1Gal—C4All—C5All) = −160.93 (10)°, where ring-atom numbering conforms to the convention in which C1 denotes the anomeric C atom, C5 the ring atom bearing the exocyclic hydroxymethyl group, and C6 the exocyclic hy­droxy­methyl (CH2OH) C atom in the βGalp and βAllp residues. Inter­nal linkage conformations in the crystal structures of the structurally related disaccharides methyl β-lactoside [methyl β-d-galactopyranosyl-(1→4)-β-d-gluco­pyran­oside] methanol solvate [Stenutz, Shang & Serianni (1999). Acta Cryst. C55, 1719–1721], (II), and methyl β-cello­bioside [methyl β-d-gluco­pyran­osyl-(1→4)-β-d-gluco­pyran­oside] methanol solvate [Ham & Williams (1970). Acta Cryst. B26, 1373–1383], (III), are characterized by φ′ = −88.4 (2)° and ψ′ = −161.3 (2)°, and φ′ = −91.1° and ψ′ = −160.7°, respectively. Inter-residue hydrogen bonding is observed between O3Glc and O5Gal/Glc in the crystal structures of (II) and (III), suggesting a role in determining their preferred linkage conformations. An analogous inter-residue hydrogen bond does not exist in (I) due to the axial orientation of O3All, yet its inter­nal linkage conformation is very similar to those of (II) and (III).

DE NOVO ASYMMETRIC APPROACH TO THE DISACCHARIDE PORTION OF SCH-47554.

A method for the asymmetric synthesis of the disaccharide portion of SCH-47554 has been developed in 6 steps. The route is shorter than the reported route to a related disaccharide. The route involves the use of the Noyori reduction to establish the asymmetry of the d- and l-sugar portion of the molecule. Diastereoselective Pd-glycosylation reaction and subsequent post-glycosylation transformation are used to establish the remaining stereocenter.

Insights into the mechanism of separation of heparin and heparan sulfate disaccharides by reverse-phase ion-pair chromatography

Reverse-phase ion-pair high performance liquid chromatography (RPIP-HPLC) and ultra-performance liquid chromatography (RPIP-UPLC) are increasingly popular chromatographic techniques for the separation of organic compounds. However, the fine details of the RPIP separation mechanism are still being debated. Many factors including type and concentration of the ion-pairing reagent, mobile phase pH, organic modifier, ionic strength, and stationary phase all play a role in the overall efficiency and optimization of ion-pairing separations. This study investigates the role that competition between ion-pairing reagents with different steric bulk and hydrophobicity plays in the separation of structural isomers of heparin and heparan sulfate (HS) disaccharides. In addition to providing insights into the mechanism by which RPIP-HPLC can resolve closely related disaccharides, the use of competition between ion-pairing agents could lead to new methods for the separation of larger heparin and HS oligosaccharides. This approach should also be applicable to the analysis of other compound classes, and could lead to a general approach for the chromatographic resolution of mixtures of charged analytes having similar structures.

Methyl β-allolactoside [methyl β-D-galactopyranosyl-(1→6)-β-D-glucopyranoside] monohydrate

Methyl beta-allolactoside [methyl beta-D-galactopyranosyl-(1-->6)-beta-D-glucopyranoside], (II), was crystallized from water as a monohydrate, C(13)H(24)O(11).H(2)O. The betaGalp and betaGlcp residues in (II) assume distorted (4)C(1) chair conformations, with the former more distorted than the latter. Linkage conformation is characterized by phi' (C2(Gal)-C1(Gal)-O1(Gal)-C6(Glc)), psi' (C1(Gal)-O1(Gal)-C6(Glc)-C5(Glc)) and omega (C4(Glc)-C5(Glc)-C6(Glc)-O1(Gal)) torsion angles of 172.9 (2), -117.9 (3) and -176.2 (2) degrees , respectively. The psi' and omega values differ significantly from those found in the crystal structure of beta-gentiobiose, (III) [Rohrer et al. (1980). Acta Cryst. B36, 650-654]. Structural comparisons of (II) with related disaccharides bound to a mutant beta-galactosidase reveal significant differences in hydroxymethyl conformation and in the degree of ring distortion of the betaGlcp residue. Structural comparisons of (II) with a DFT-optimized structure, (II(C)), suggest a link between hydrogen bonding, pyranosyl ring deformation and linkage conformation.

Progress in biomimetic carbohydrate recognition.

The importance of carbohydrate recognition in biology, and the unusual challenges involved, have lead to great interest in mimicking saccharide-binding proteins such as lectins. In this review, we discuss the design of artificial carbohydrate receptors, focusing on those which work under natural (i.e. aqueous) conditions. The problem is intrinsically difficult because of the similarity between substrate (carbohydrate) and solvent (water) and, accordingly, progress has been slow. However, recent developments suggest that solutions can be found. In particular, the "temple" family of carbohydrate receptors show good affinities and excellent selectivities for certain all-equatorial substrates. One example is selective for O-linked beta-N-acetylglucosamine (GlcNAc, as in the O-GlcNAc protein modification), while another is specific for beta-cellobiosyl and closely related disaccharides. Both show roughly millimolar affinities, matching the strength of some lectin-carbohydrate interactions.

Conversion of mono/di/polysaccharides into furan compounds using 1-alkyl-3-methylimidazolium ionic liquids

The one-pot hydrolysis/dehydration of mono/di/polysaccharides into furfural (FUR) or 5-hydroxymethyl-2-furaldehyde (HMF) in the presence of an acidic ionic liquid, 1-ethyl-3-methylimidazolium hydrogen sulfate ([EMIM][HSO 4]), at 100 °C, was investigated and the results were compared with those using 1-butyl-3-methylimidazolium chloride ([BMIM][Cl]), with or without CrCl 3 or H 2SO 4. The ionic liquid [EMIM][HSO 4] is effective in converting xylose and fructose or related polysaccharides into FUR (e.g. from xylose: 84% yield at 6 h) or HMF (e.g. from fructose: 88% yield at 30 min), but not glucose and polymers containing these units, which contrasts to that observed for the [BMIM][Cl]/CrCl 3 system. The latter is quite effective in converting glucose and related disaccharides into HMF, but not the polysaccharides cellulose and starch. For the latter feedstocks, the addition of H 2SO 4 or [EMIM][HSO 4] to cellulose/[BMIM][Cl]/CrCl 3 enhances HMF yields, presumably by accelerating the hydrolysis step, although the selectivity of the dehydration of glucose monomers seems rather poor (HMF yield < 13 wt.%). In the case of the xylose to FUR conversion in [EMIM][HSO 4], two systems involving the reaction and simultaneous separation of the target product were investigated, namely reaction/solvent extraction and reaction/evaporation systems, and for both the ionic liquid could be recovered and reused without a significant drop in FUR yield in recycling runs.

PapA3 Is an Acyltransferase Required for Polyacyltrehalose Biosynthesis in Mycobacterium tuberculosis

Mycobacterium tuberculosis possesses an unusual cell wall that is replete with virulence-enhancing lipids. One cell wall molecule unique to pathogenic M. tuberculosis is polyacyltrehalose (PAT), a pentaacylated, trehalose-based glycolipid. Little is known about the biosynthesis of PAT, although its biosynthetic gene cluster has been identified and found to resemble that of the better studied M. tuberculosis cell wall component sulfolipid-1. In this study, we sought to elucidate the function of papA3, a gene from the PAT locus encoding a putative acyltransferase. To determine whether PapA3 participates in PAT assembly, we expressed the protein heterologously and evaluated its acyltransferase activity in vitro. The purified enzyme catalyzed the sequential esterification of trehalose with two palmitoyl groups, generating a diacylated product similar to the 2,3-diacyltrehalose glycolipids of M. tuberculosis. Notably, PapA3 was selective for trehalose; no activity was observed with other structurally related disaccharides. Disruption of the papA3 gene from M. tuberculosis resulted in the loss of PAT from bacterial lipid extracts. Complementation of the mutant strain restored PAT production, demonstrating that PapA3 is essential for the biosynthesis of this glycolipid in vivo. Furthermore, we determined that the PAT biosynthetic machinery has no cross-talk with that for sulfolipid-1 despite their related structures.

Sucrose metabolism contributes to in vivo fitness of Streptococcus pneumoniae.

We characterized two sucrose-metabolizing systems -sus and scr- and describe their roles in the physiology and virulence of Streptococcus pneumoniae in murine models of carriage and pneumonia. The sus and scr systems are regulated by LacI family repressors SusR and ScrR respectively. SusR regulates an adjacent ABC transporter (susT1/susT2/susX) and sucrose-6-phosphate (S-6-P) hydrolase (susH). ScrR controls an adjacent PTS transporter (scrT), fructokinase (scrK) and second S-6-P hydrolase (scrH). sus and scr play niche-specific roles in virulence. The susH and sus locus mutants are attenuated in the lung, but dispensable in nasopharyngeal carriage. Conversely, the scrH and scr locus mutants, while dispensable in the lung, are attenuated for nasopharyngeal colonization. The scrH/susH double mutant is more attenuated than scrH in the nasopharynx, indicating SusH can substitute in this niche. Both systems are sucrose-inducible, with ScrH being the major in vitro hydrolase. The scrH/susH mutant does not grow on sucrose indicating that sus and scr are the only sucrose-metabolizing systems in S. pneumoniae. We propose a model describing hierarchical regulation of the scr and sus systems by the putative inducer, S-6-P. The transport and metabolism of sucrose or a related disaccharide thus contributes to S. pneumoniae colonization and disease.

Polyvalent GalNAcα1→Ser/Thr (Tn) and Galβ1→3GalNAcα1→Ser/Thr (T α) as the most potent recognition factors involved in Maclura pomifera agglutinin–glycan interactions

The agglutinin isolated from the seeds of Maclura pomifera (MPA) recognizes a mucin-type disaccharide sequence, Galbeta1-->3GalNAc (T) on a human erythrocyte membrane. We have utilized the enzyme-linked lectinosorbent assay (ELLSA) and inhibition assay to more systematically analyze the carbohydrate specificity of MPA with glyco-recognition factors and mammalian Gal/GalNAc structural units in lectin-glycoform interactions. From the results, it is concluded that the high densities of polyvalent GalNAcalpha1-->Ser/Thr (Tn) and Galbeta1-->3GalNAcalpha1-->Ser/Thr (T(alpha)) glycotopes in macromolecules are the most critical factors for MPA binding, being on a nanogram basis 2.0 x 10(5), 4.6 x 10(4) and 3.9 x 10(4) more active than monovalent Gal, monomeric T and Tn glycotope, respectively. Other carbohydrate structural units in mammalian glycoconjugates, such as human blood group Sd (a+) related disaccharide (GalNAcbeta1-->4Gal) and Pk/P1 active disaccharide (Galalpha1-->4Gal) were inactive. These results demonstrate that the configurations of carbon-4 and carbon-2 are essential for MPA binding and establish the importance of affinity enhancement by high-density polyvalencies of Tn/T glycotopes in MPA-glycan interactions. The overall binding profile of MPA can be defined in decreasing order as high density of polyvalent Tn/T(alpha) (M.W. > 4.0 x 10(4)) >> Tn-containing glycopeptides (M.W. < 3.0 x 10(3)) > monomeric T/Tn and P (GalNAcbeta1-->3Gal) > GalNAc > Gal >> Man, L: ARA: , D: Fuc and Glc (inactive). Our findings should aid in the selection of this lectin for elucidating functions of carbohydrate chains in life processes and for applications in the biomedical sciences.