Thermodynamic and kinetic studies on the mechanism of binding of methylumbelliferyl glycosides to jacalin

The binding of Artocarpus integrifolia lectin (jacalin) to 4-methylumbelliferyl (Meumb)-glycosides, Gal alpha Meumb, Gal beta Meumb, GalNAc alpha Meumb, GalNAc beta-Meumb, and Gal beta 3GalNAc beta Meumb was examined by extrinsic fluorescence quenching titration and stopped flow spectrofluorimetry. The binding was characterized by 100% quenching of fluorescence of Meumb-glycosides. Their association constants range from 2.0 x 10(4) to 1.58 x 10(6) M-1 at 15 degrees C. Entropic contribution is the major stabilizing force for avid binding of Meumb-glycosides indicating the existence of a hydrophobic site that is complementary to their methylumbelliferyl group. The second order association rate constants for interaction of these sugars with lectin at 15 degrees C vary from 8.8 x 10(5) to 3.24 x 10(6) M-1 S-1, at pH 7.2. The first order dissociation rate constants range from 2.30 to 43.0 S-1 at 15 degrees C. Despite the differences in their association rate constants, the overall values of association constants for these saccharides are determined by their dissociation rate constants. The second order rate constant for the association of Meumb-glycosides follows a pattern consistent with the magnitude of the activation energies involved therin. Activation parameters for association of all ligands illustrate that the origin of the barrier between binding of jacalin to Meumb-glycosides is entropic, and the enthalpic contribution is small. A correlation between these parameters and the structure of the ligands on the association rates underscores the importance of steric factors in determining protein saccharide recognitions.

Role of hydroxyl group on the mesomorphism of alkyl glycosides:synthesis and thermal behavior of alkyl 6-deoxy-beta-D-glucopyranosides

A homologous series of alkyl 6-deoxy-beta-D-glucopyranoside amphiphiles was prepared,in an effort to identify the role of hydroxyl group in the mesomorphic behavior of alkyl glycosides. Synthesis was performed by a chlorination of the sugar moiety in alkyl-beta-D-glucopyranosides with methylsulfonyl chloride in DMF, followed by a metal mediated dehalogenation to secure alkyl 6-deoxy-beta-D-glucopyranosides, wherein the alkyl chain length varied from C-9 to C-16. The mesomorphic behavior of these 6-deoxy alkyl glycosides was assessed using polarizing optical microscopy, differential scanning calorimetry and X-ray diffraction method. Whereas the lower homologues exhibited a monotropic SmA phase till sub-ambient temperatures, the higher homologues formed a plastic phase. A partial interdigitized bilaye structure of SmA phase is inferred from experimental d-spacing and computationally derived lengths of the molecules. The results were compared with those of normal alkyl glucopyranosides, retained with hydroxyl groups at C-2-C-6 carbons, and alkyl 2-deoxy-glucopyranosides, devoid of a hydroxyl group at C-2 and the comparison showed important differences in the mesomorphic behavior.(C)2010 Elsevier Ireland Ltd. All rights reserved.

Crystal structures and thermal analyses of alkyl 2-deoxy-alpha-D-arabino-hexopyranosides

The crystal structures of alkyl 2-deoxy-alpha-D-arabino-hexopyranosides, with the alkyl chain lengths from C-8 to C-18, are established by the single crystal X-ray structural determination. The even-alkyl chain length derivatives crystallized orthorhombic, with space group P2(1)2(1)2(1), whereas the odd-alkyl chain length derivatives crystallized monoclinic, with space group P2(1). The sugar moieties retained a C-4(1) chair conformation and the conformation of the alkyl chains was all-trans. The molecules formed a bilayer structure, in which alkyl chains were interdigitated.The hydrogen bonds, originating from the sugar moieties, were observed in adjacent layers and also within the same layer, resulting in the formation of infinite chains. The alkyl chains arranged parallel to each other and formed planar structures. The thermal properties of the alkyl 2-deoxy glucosides were analyzed further. It was observed that none of the derivatives exhibited mesomorphism. This study establishes that the absence of the hydroxyl group at C-2 of the sugar moiety results in a non-mesogenic nature of the alkyl 2-deoxy-alpha-D-glycosides, as opposed to the profound mesogenic nature of the normal alkyl glycosides.

Reactivity switching and selective activation of C-1 or C-3 in 2,3-unsaturated thioglycosides

Reactivity switching and selective activation of C-1 or C-3 in 2,3-unsaturated thioglycosides, namely, 2,3-dideoxy-1-thio-D-hex-2-enopyranosides are reported. The reactivity switching allowed activation of either C-1 or C-3, with the use of either N-iodosuccinimide (NIS)/triflic acid (TfOH) or TfOH alone. C-1 glycosylation with alcohol acceptors occurred in the presence of NIS/TfOH, without the acceptors reacting at C-3. On the other hand, reaction of 2,3-unsaturated thioglycosides with alcohols mediated by triflic acid led to transposition of C-1 ethylthio-moiety to C-3 intramolecularly, to form 3-ethylthio-glycals. Resulting glycals underwent glycosylation with alcohols to afford 3-ethylthio-2-deoxy glycosides. However, when thiol was used as an acceptor, only a stereoselective addition at C-3 resulted, so as to form C-1, C-3 dithio-substituted 2-deoxypyranosides. (C) 2011 Elsevier Ltd. All rights reserved.

Increased glycosidic bond stabilities in 4-C-hydroxymethyl linked disaccharides

Three new hydroxymethyl-linked non-natural disaccharide analogues, containing an additional methylene group in between the glycosidic linkage, were synthesized by utilizing 4-C-hydroxymethyl-alpha-D-glucopyranoside as the glycosyl donor. A kinetic study was undertaken to assess the hydrolytic stabilities of these new disaccharide analogues toward acid-catalyzed hydrolysis, at 60 degrees C and 70 degrees C. The studies showed that the disaccharide analogues were stable, by an order of magnitude, than naturally-occurring disaccharides, such as, cellobiose, lactose, and maltose. The first order rate constants were lower than that of methyl glycosides and the trend of hydrolysis rate constants followed that of naturally-occurring disaccharides. alpha-Anomer showed faster hydrolysis than the beta-anomer and the presence of axial hydroxyl group also led to faster hydrolysis among the disaccharide analogues. Energy minimized structures, derived through molecular modeling, showed that dihedral angles around the glycosidic bond in disaccharide analogues were nearly similar to that of naturally-occurring disaccharides. (C) 2011 Elsevier Ltd. All rights reserved.

Synthesis of 2-Deoxy-2-C-alkyl Glycal and Glycopyranosides from 2-Hydroxy Glycal Ester

A method to convert 2-hydroxy glycol ester to the corresponding corresponding 2-deoxy-2-C-alkyl glycol in a facile manner, through key reactions including (i) C-allylation at C-1, (ii) Wittig reaction, and (iii) Cope rearrangement of a 1,5-diene derivative, is reported. The alpha-anomer of the 1,5-diene derivative underwent Cope rearrangement to afford 2-deoxy-2-C-glycal derivative, whereas the beta-anomer was found to be unreactive. Employing this sequence, was transformed to 3,4,6-tri-O-benzyl-2-deoxy-2-C-alkyl-1,5-anhydro-D-arabino-hex-1-enitol. 2-Deoxy-2-C-alkyl glycol derivative is a suitable glycosyl donor to prepare 2-deoxy-2-C-alkyl glycosides, mediated through haloglycosylation and a subsequent dehalogenation. A number of 2-deoxy-2-C-alkyl glycosides, with both glycosyl and nonglycosyl moieties at the reducing end, are thus prepared from the glycol.

Functional characterization, homology modeling and docking studies of beta-glucosidase responsible for bioactivation of cyanogenic hydroxynitrile glucosides from Leucaena leucocephala (subabul)

Glycosyl hydrolase family 1 beta-glucosidases are important enzymes that serve many diverse functions in plants including defense, whereby hydrolyzing the defensive compounds such as hydroxynitrile glucosides. A hydroxynitrile glucoside cleaving beta-glucosidase gene (Llbglu1) was isolated from Leucaena leucocephala, cloned into pET-28a (+) and expressed in E. coli BL21 (DE3) cells. The recombinant enzyme was purified by Ni-NTA affinity chromatography. The optimal temperature and pH for this beta-glucosidase were found to be 45 A degrees C and 4.8, respectively. The purified Llbglu1 enzyme hydrolyzed the synthetic glycosides, pNPGlucoside (pNPGlc) and pNPGalactoside (pNPGal). Also, the enzyme hydrolyzed amygdalin, a hydroxynitrile glycoside and a few of the tested flavonoid and isoflavonoid glucosides. The kinetic parameters K (m) and V (max) were found to be 38.59 mu M and 0.8237 mu M/mg/min for pNPGlc, whereas for pNPGal the values were observed as 1845 mu M and 0.1037 mu M/mg/min. In the present study, a three dimensional (3D) model of the Llbglu1 was built by MODELLER software to find out the substrate binding sites and the quality of the model was examined using the program PROCHEK. Docking studies indicated that conserved active site residues are Glu 199, Glu 413, His 153, Asn 198, Val 270, Asn 340, and Trp 462. Docking of rhodiocyanoside A with the modeled Llbglu1 resulted in a binding with free energy change (Delta G) of -5.52 kcal/mol on which basis rhodiocyanoside A could be considered as a potential substrate.