Raymond U. Lemieux

The conformational properties of sucrose in aqueous solution: intramolecular hydrogen-bonding

Abstract A detailed analysis is presented of the nuclear ( 1 H and 13 C) magnetic resonance (n.m.r.) properties of sucrose, using both D 2 O and dimethyl sulfoxide- d 6 as solvents, based on measurements of coupling constants, chemical shifts, T 1 relaxation times, and nuclear Overhauser enhancements. Molecular modelling (HSEA calculations) suggests a strong conformational preference about the glycosidic linkages that is near to that for sucrose in the crystalline state, and this conformational rigidity is fully supported by the n.m.r. data, in terms of lack of influence of changes in concentration and temperature on the relevant n.m.r. parameters. The restricted rotation for the 1-hydroxymethyl group of the fructose residue is related to the persistence of the intramolecular hydrogen-bond between O-1 f and O-2 g . The presence of this bond was established for solutions in (CD 3 ) 2 SO by the observation of isotopic chemicashifts on partial deuteration of the hydroxyl groups. The orientation of the 6-hydroxyl methyl group of the fructose residue is not that present in the crystalline state but, in (CD 3 ) 2 SO, it may be intramolecularly hydrogen-bonded, as was demonstrated by titration of the hydroxyl groups with CD 3 OD. Observations are made regarding hydrophobic topographies common to sucrose, saccharin, and 1-chloro-1-deoxysucrose, which may have a bearing on sweetness.

Applications of empirical rules for optical rotation to problems of conformational analysis

Abstract Whiffens rules for estimating the molar rotation of saturated pyranoid carbohydrates from their geometrical structure are reduced to four rotational parameters, based on gauche relationships between carbon atoms, oxygen atoms, and carbon and oxygen atoms, and whether or not these atoms are bridged by C-C or O-C. The numerical values assigned are based on the rotations of simple model compounds as structural units of the more complex molecules. Consideration is given to the favored orientations for the methyl group of methyl glycopyranosides and for the methyl group of hexopyranoses and their derivatives. Conclusions based on rotation are substantiated by the coupling of 13 C to the anomeric hydrogen atom for the methyl glycopyranosides. Solvation effects on rotation are interpreted in terms of the empirical rules and the effects of hydrogen bonding.

The conformational analysis of oligosaccharides by 1H-NMR and HSEA calculation

The application of 1H-nuclear Overhauser enhancement, 1H-spin-lattice-relaxation-time and 1H-chemical shift measurements for the assessment of the conformational preferences of oligosaccharides are briefly reviewed. It is demonstrated that additivity rules, for the correlation of the chemical shifts of similar hydrogen atoms in different oligosaccharides, can be useful in the conformational analysis of oligosaccharides when the differential chemical shifts are greater than 0.1 ppm. These often can be attributed to specific interunit deshielding of a hydrogen atom by an oxygen atom with which it is in strong nonbonded interaction. HSEA calculations are used to demonstrate that differential chemical shifts of less than 0.1 ppm can have origins that are not significant to the overall conformational preferences of the oligosaccharides which are being compared. Both shielding and deshielding effects can arise from a change in the orientation of a substituent group as the result of the introduction of a sugar on a neighboring unit. It is demonstrated that substituent groups, such as hydroxymethyl and acetamido groups, on occasions, should be treated in HSEA calculations as freely rotating about their linkage to a pyranose ring.

Synthetic, conformational, and immunochemical studies of modified Lewis b and Y human blood-group determinants to serve as probes for the combining site of the lectin IV of Griffonia simplicifolia

Syntheses of the methyl glycosides of the Lewis b [alpha-L-Fuc-(1----2)-beta-D-Gal-(1----3) [alpha-L-Fuc-(1----4)]-beta-D-GlcNAc-] and Y [alpha-L-Fuc-(1----2)-beta-D-Gal-(1----4) [alpha-L-Fuc-(1----3)]-beta-D- GlcNAc-] human blood-group determinants and both their 6a-deoxy and N-deacetylated derivatives are reported. In the case of the Lewis b structure (Leb-OMe), the 6a-O-mesyl and 6a-deoxy-6a-iodo derivatives were also prepared. The conformational preferences predicted by HSEA calculation are shown to be in good agreement with expectations based on 1H- and 13C-n.m.r. spectroscopy. The immunochemical data based on inhibition and thermodynamic studies require that the binding of Leb-OMe and Y-OMe by the lectin IV of Griffonia simplicifolia does not involve recognition of the OMe, NHAc, or 6a-OH group and, consequently, occurs at a cleft at the surface of the protein. The complex formed between the lectin and 6a-deoxy-6a-iodo-Leb-OMe provided the heavy nuclei required for the solution of the X-ray crystal structure.

Synthesis of the T [β-d-Gal-(1→3)-α-d-GalNAc]-antigenic determinant in a form useful for the preparation of an effective artificial antigen and the corresponding immunoadsorbent

Abstract The T antigenic determinant was synthesized in the form 8-methoxycarbonyloctyl 2-acetamido-2-deoxy-3- O -(β- d -galactopyranosyl)-α- d -galactopyranoside ( 6 ) (β- d -Gal-(1→3)-α- d -GalNAcO(CH 2 ) 8 CO 2 Me). This T-hapten was used to prepare a T-BSA artificial antigen ( 7 ) and an immunoadsorbent ( 8 ), which were shown to possess the expected immunological properties. Nuclear Overhauser enhancements of the signals for anti -periplanar H-2′ and the syn -axial H-3′ of the β- d -galactopyranosyl group were observed on saturation of H-1′. The signal for H-3 of the 2-acetamido-2-deoxy-α- d -galactopyranoside residue was also enhanced.

The synthesis of monodeoxy derivatives of lacto-N-biose I and N-acetyl-lactosamine to serve as substrates for the differentiation of α-l-fucosyl transferases

Abstract The chemical synthesis of β- d -Gal-(1→3)-4-deoxy-β- d -GlcNAcOR and β- d -Gal-(1→4)-3-deoxy-β- d -GlcNAcOR, wherein R = (CH2)8COOMe and Me, are reported. Also, 2-deoxy-β- d -Gal-(1→3)-β- d -GlcNAcO(CH2)8COOMe and 2-deoxy-β- d -Gal-(1→4)-β- d -GlcNAcO(CH2)8 COOMe were synthesized. Preliminary results from other laboratories are reported on the behaviour of these compounds as acceptors of β- d -Gal- and β- d -GlcNAc-α- l -fucosyl transferases. It is suggested that the enzymes accept the substrates in near their most favorable conformations and that the binding involves both polar and nonpolar interactions with topographical features near the hydroxyl group which undergoes substitution. It appears that, depending on the transferase, the recognition may or may not include both the sugar units.

The recognition of three different epitopes for the H-type 2 human blood group determinant by lections ofUlex europaeus, Galactia tenuiflora andPsophocarpus tetragonolobus (Winged Bean)

The chemical mapping of the regions of H-type 2 human blood group-related trisaccharide (Fucα(1–2)Galβ(1–4)GlcNAcβMe) that are recognized by three different lectins, the so-called epitopes, are reviewed together with an account of how and why oligosaccharides form specific complexes with proteins as presently viewed in this laboratory. The occasion is used to report the synthesis of the various mono-O-methyl derivatives of the above trisaccharide that were used in these investigations. Also, Fucα(1–2)Galβ(1–4)XylβMe was synthesized in order to examine whether or not the hydroxymethyl group of the GlcNAc residue participates in the binding reaction.

Immunochemical studies on blood groups—LXIX. The conformation of the trisaccharide determinant in the combining site of anti-I Ma (group 1)

Abstract Further studies of the inhibition of the monoclonal anti-I Ma (group 1) antibody, in quantitative precipitin assays, by synthetic oligosaccharides derived from N-acetyllactosamine (LacNAc) are reported. The results confirm that the antibody binds about the βLacNAc-OCH2CHO-portions of such structures as βLacNAc(1→6)β d Gal or βLacNAc(1→6)α d GalNAc. The trisaccharides βLacNAc(1→6)-7-deoxy- d -glycero- d -galacto-heptose (7) and βLacNAc(1→6)-7-deoxy- l -glycero- d -galacto-heptose (8), in contrast to βLacNAc(1→6) d Gal (1), were found (1HNMR) to possess well-defined conformations. Since 7 proved to be a 2.2 times better inhibitor than 1, whereas 8 was a poor inhibitor, it is concluded that the antibody binds 1 in the conformation which is preferred by 7. On this basis, it is noted that 1 would reside in a conformation that possesses a region amenable to hydrophobic bonding that extends between the C-5 groupings of the two d Gal units and over the acetamido group of the central β d GlcNAc residue with the 3-hydroxyl of this unit hydrogen bonded to O-5 of the terminal β d Gal unit. It is suggested that it is this region which becomes bound to a hydrophobic site of the antibody. Thus, nine of the hydroxyl groups in 1 would remain available for interaction with the solvent water and it would follow that the combining site is an hydrophobic cleft.

Competition between ABO and Le gene specified enzymes

Abstract. Radioimmunoassays were prepared using two anti‐A and one anti‐B reagents. The specificity of the procedures was assessed with 13 artificial antigens. The amounts of A and B natural antigens in saliva of ABH secretors of known Lewis phenotype were measured with these assays. The results confirmed that the average amount of A antigen is lower in Lewis‐positive (Leb) than in Lewis‐negative (Led) donors and in A2 than in A1 donors. However, the differences among the four combined A and Lewis phenotypes were only supported by significantly lower amounts of A antigenic determinants in A2Leb as compared to the other three phenotypes (A1 Leb, A1Led and A2Led) that had similar amounts of A antigenic determinants. No Lewis‐related difference could be detected in the amounts of B antigens between BLeb and BLed donors. The results are discussed in terms of competition between A, B and Lewis‐gene‐specified enzymes for their common acceptors. The difference in the efficiency of the A2 enzyme as compared to that of the A1 enzyme is proposed as a possible explanation for the A1‐A2 phenotypic difference.

The binding of the Lewis-a human blood group determinant by two hybridoma monoclonal anti-Lea antibodies

The combining sites of two hybridoma monoclonal antibodies with specificities for the Lewis-a human blood group determinant, beta-D-Galp-(1----3)-[alpha-L-Fucp-(1----4)]-beta-D-GlcpNAc-(1---- , were probed by use of a wide range of inhibitors of the reaction between an 125I-labelled artificial Lewis-a antigen and the antibodies under the conditions of a solid-phase radioimmunoassay. Amongst the inhibitors examined were the eight possible monodeoxy derivatives of the trisaccharide methyl glycoside. As was previously found for other antibodies and lectins, the results indicated that, in each case, a cluster of polar groups provide a key polar interaction with the protein. The further involvement of large lipophilic regions of the trisaccharide was also indicated. These regions are expected to provide an essentially nonpolar interaction for complex formation, and appear to include intramolecularly hydrogen-bonded hydroxyl groups. Although the features of the trisaccharide that are recognized are similar in kind, the patterns are very different. For example, in the case of antibody AH8-34, the key polar group is provided by OH-2 and OH-3 of the beta-D-Galp group, whereas for the other antibody (CF4-C4) OH-3 and OH-4 of the beta-D-Galp group and OH-4 of the alpha-L-Fucp group provide the key polar interaction. It appears that, for AH8-34, the trisaccharide is accepted with OH-2 of the beta-D-Galp group and OH-3 of the alpha-L-Fucp group intramolecularly hydrogen-bonded to neighboring proton acceptors. For CF4-C4, such intramolecular hydrogen-bonding appears to involve OH-4 and OH-6 of the beta-D-Galp group.