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Oligosaccharide structures: theory versus experiment
617
Oligosaccharide structures: theory versus experiment Anne Imberty Recently, the interdependency of theoretical and experimental approaches in the structure determination of oligosaccharides has been confirmed. More accurate simulations are possible because of the advances in software and computers. Meanwhile, improvements in NMR techniques permit the measurement of numerous structural and dynamical parameters, either for the free state or for carbohydrate ligands bound to receptors. Several crystal structures of isolated or protein-complexed oligosaccharides give new clues for modeling the intermolecular forces that drive the interactions.
Addresses Centre de Recherches sur les Macromolacules Veg6tales, Joseph Fourier Universite, BP53, F38041, Grenoble cedex 9, France; e-mail: [email protected] Current Opinion in Structural Biology 1997, 7:61 ?-623 http://biomednet.com/elecref/O959440X0070061 ? © Current Biology Ltd ISSN 0959-440X Abbreviations MBP mannose-binding protein MD molecular dynamics NOE nuclear Overhauser effect sLe x sialyl Lewisx
Introduction In many instances, even extensive knowledge of the primary structure of oligosaccharides is insufficient to understand and explain their function and specificity. T h e 3D structures of glycans provide the driving force for all intermolecular interactions and therefore predetermine their function; furthermore, the flexibility and dynamics of oligosaccharides frequently play a key role in biological activity and must also be taken into account. In recent years, it has been recognized that efficient carbohydrate modeling demands the use of appropriate force-fields, as conformational preference at the glycosidic linkage is due to special stereoelectronic effects. Some of the general force-fields, such as MM3, have proven to be suitable for this purpose, whereas others require supplementary parametrization. During the past two years, several new parametrizations for carbohydrates have been proposed for the programs more widely used in the domain of molecular dynamics (MD). Examples can be found for AMBER [1-3], C H A R M m [41 and GROMOS [5]. As a result of theses advances, some extended MD simulations have been routinely calculated for disaccharides [6], and the inclusion of explicit water molecules has been facilitated [7",8°°].
Even if modeling groups are generally aware of the conformational particularities of carbohydrates, one must keep in mind the fact that any model proposed via a theoretical study should, where possible, be validated by a comparison with experimental data. X-ray crystallography provides the most precise information on the shape of a carbohydrate molecule. In addition, a thorough analysis of packing arrangements frequently produces information on preferential carbohydrate-carbohydrate and carbohydrate-water relationships. A limitation of the above method is that it does not reflect oligosaccharide flexibility. A more fundamental point to be considered is the difficulty encountered in the crystallization of oligosaccharides and even disaccharides. N M R experiments are widely used for the validation of structures of oligosaccharides using predicted molecular modeling. Measurements of nuclear Overhauser effects (NOEs) help in the determination of conformations in solution and, in addition, have the tremendous advantage of giving access to some dynamic properties of molecules such as the timescales of internal motions. During the past two years, the major advances have been in the determination of the bioactive conformation of oligosaccharides, that is, their conformation when bound to a protein receptor. In solution, transferred N O E experiments have proven to be a powerful tool in the investigation of the bound conformations of oligosaccharides [9",10°°]. In the past year, this method has been successfully used as a tool for the conformational analysis of bound carbohydrates and analogs [I 1",12,13,14°°,15°,16°]. In the mean time, several new crystal structures of plant lectin-oligosaccharide complexes have been solved [17-19,20°]. For some linkages, such as those of the Manc~(1-3)Man and GIcNAc[3(1-2)Man disaccharides, crystal structures of lectin-oligosaccharide complexes provide sufficient evidence to illustrate that the conformations selected upon binding indeed correspond to the several energy minima predicted by theoretical calculations performed on individual glycosidic linkages (Figure 1). I will review several recent examples of oligosaccharide structure for which both theoretical and experimental data are available.
Comparison of the conformation of lactose and its C-analog T h e conformational behavior of the lactose disaccharide (]3Gal[1-4]GIc) has been well characterized using a combination of theoretical and experimental methods. Its potential energy surface is characteristic of equatorial-equatorial (1-4)-linked disaccharides possessing three or four major potential-energy wells. T h e most populated conformation
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is the so-called syn, which corresponds to all observed conformations of lactose crystal structures, including those found in protein complexes. Anti and gauche-gauche conformations differ from the lowest energy shape by a large change in the ~) and gt torsion angles, respectively [21,22]. As each of the three conformations has a characteristic NOE fingerprint (Figure 2), it has been possible to deduce that the population of lactose in solution is about 90% svn and 10% anti, with no detectable gauche-gauche conformer [14"*,21]. Because of its well-characterized conformational behavior, lactose is now used for testing force-fields [22,23"]. Recentl'> C-disaccharides have been investigated as potential mimetics of natural disaccharides. As the glycosidic oxygen is replaced by a methylene group, these compounds exhibit a higher chemical and biochemical stability. A significant interest has arisen in comparing the molecular mechanics and dynamics behavior of C-disaccharide with those of O-disaccharide to determine whether such compounds can act as competitive inhibitors of their O-counterparts. A conformational study of C-lactose using a combination of NMR spectroscopy and molecular mechanics has shown that it is more flexible than lactose [24]. Three main low-energy conformations ate predicted to coexist in solution with a distribution of 55%, 40%, and 5% for syn, anti, and gauche-gauche, respectively. Another study has corroborated the flexibility of C-lactose [25"],
but the comparison of experimental and back-calculated N O E values predicts the presence of only two conformers, namely syn (60%) and anti (40%). Both O-lactose and C-lactose conformations have also been studied in the protein-bound state. When methyl [3-1actoside [26] or methyl o~-lactoside [27] are bound to ricin B, the measured transferred N O E implies only a slight variation from the lowest energy anti conformation. T h e situation is completely different with C-lactose. In the presence of ricin B, only the transferred NOE characteristic of the syn conformation can be observed [13,14"]. These experimental results indicate, therefore, that ricin B selects different conformers of C-lactose and O-lactose. Thus, it appears that it is possible for the inhibitor conformation selected by the protein receptor to be different from that of the natural substrate. Such conformational behavior could prove to be vital when designing new oligosaccharide mimics. Histo-blood group oligosaccharides Conformational analysis of histo-blood group carbohydrate determinants was pioneered by Lemieux et al. [28] and Rao and Biswas [29] more than 15 years ago (Figure 3). These molecules have been the subject of much interest in recent years, while the role of the Lewis determinants has been discovered in many biological functions. Conformational analysis of all constituting disaccharides
Oligosaccharide structures Imberty
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Graphic representation of the three low-energy conformations of (x-methyl lactose. (a) Syn. (b) Anti. (c) Gauche-gauche. The characteristic NOE signature of each conformer is indicated by arrows.
of blood group oligosaccharides has been performed recently [30,31]. For comparison, the only disaccharide fragments for which crystal structures are available arc N-acetyllactosamine [32], which was crystallized more than 10 years ago, and the H-type disaccharide otFuc(1-2)13Gal (blood group O determinant) [33]. In both cases, the crystal conformation corresponds to the predicted lowest energy regions. All the constituent disaccharides are predicted to be flexible. T h e conformational behavior of histo-blood group oligosaccharides is different. Depending on their branch-
ing patterns, they can exhibit a range of conformational behavior, from almost complete rigidity to high flexibility [30,34,35]. T h e different alignments of oligosaccharides that have been deduced from these studies have been used to determine the epitopes recognized by monoclonal antibodies [36]. In a more in-depth study, 3D quantitative structure/activity relationship (QSAR) methods have been applied to these molecules in order to understand the cross-reaction patterns of antiblood group antibodies [37°]. In this case, the agreement of molecular mechanics models with immunological data represents an experimental validation of the theoretical study.
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The Lewis x trisaccharide
The Lewis x determinant, [3Gal(1-4)lotFuc(1-3)]13GlcNAc, is a trisaccharidc of biological interest as it is a stagespecific embryonic marker involved in cell-cell interactions and a signaling molecule for some host-pathogen recognition. All the theoretical studies [28-31] have predicted one principal low-energy conformation, characterized by a stacking between the fucosc and galactose rings. The otFuc(1-3)13GlcNAc linkage conformation corresponds to one of the two lowest energy minima for the disaccharide, whereas the fucose-galactose interaction shifts the I3Gal(1-4)I3GlcNAc linkage to the border of a conformational plateau. Other conformational families only possibly exist at significantly higher energies; however, they can occur in special cases, such as interactions with a receptor. The conformation predicted from theoretical studies has been fully confirmed by NMR experiments on the Lewis x trisaccharide [38,39].
otGalNAc(1-3)[otFuc(1-2)]13Gal, two main conformations are predicted to occur, which differ mainly by the orientation of the fucose residue. Comparison of the simulated build-up NOE curves with experimental ones [11"] permits the confirmation of the presence of an equilibrium between these two conformers. Interaction of blood group A determinant with the lcctin from Dolichos biflorus seeds has been characterized by negative transferred NOEs corresponding only to one conformation of the trisaccharide. Theoretical transferred NOEs, calculated using an extended procedure for the complete relaxation matrix analysis of multispin exchanging systems [46], have confirmed that only one conformation is selected for binding the lectin. This result is not obvious from molecular modeling alone, as both solution conformations can be satisfactorily docked in the binding site of the D. biflorus lectin. Sialyl Lewis x
Interestingly, NMR studies performed on analogs have demonstrated a larger flexibility. A thorough 1H NMR relaxation study coupled with MD simulation [40,41 °] performed on otGalNAc(1-3)[3Gal-(1-4)[o~Fuc(1-3)]13GlcOMe has indicated that the fucosc is more mobile, presumably because of the lack of an acetamido group on the reducing glucose. The same conclusions have been reached for an analog in which the O-glycosidic bond linking fucose to glucose has been replaced by a C-glycosidic bond [42"]. In this case, longitudinal and transverse dipolar cross-relaxation rates have been measured with great accuracy, allowing a direct derivation of both geometrical and dynamical parameters. From a conformational point of vie~, therefore, studying the crystal structure of such a rigid compound may seem pointless. Nevertheless, some very interesting points have arisen from the resolution of the crystal structure [43"',44]. First, the comparison of the two independent molecules contained in the asymmetric unit has indicated that both adopt the global low-energy conformation, but with variations of - 1 0 ° in the torsion angles of the glycosidic linkages thereby confirming the local flexibility predicted by modeling and NMR studies. Second, the structure contains nine water molecules in the asymmetric unit, which is very unusual in a carbohydrate crystal structure. Lastly, the interactions between adjacent trisaccharides that are observed in the crystal packing correspond to LcX-Le x interactions that could bc involved in the formations of glycosidic clusters at the surfaces of cells and in carbohydrate-mediated cell-cell interactions [45]. The crystal structure is, therefore, a possible template for further modeling of carbohydrate-carbohydrate interactions. The blood group A determinant
Initially described as a rigid molecule [28,29], blood group A determinant has now been proposed to have several possible conformations, as predicted by a molecular mechanics study [30]. For the blood group A trisaccharide
Sialyl Lewis x (sLe x) has been the subject of intensive structural studies during the past few years because of its involvement in cellular adhesion. Molecular modeling studies agree in regard to the description of the tetrasaccharide being a rigid core consisting of the Lewis x moiety attached to sialic acid via a rather flexible linkage--two or three conformations being accessible to the molecule [34,38,47]. NMR studies with NOE measurements of the free tetrasaccharide all agree on the flexibility of the sialic acid moiety [16°,48,49]. The same conclusion has been derived from a NMR spectroscopy and molecular dynamics study of sialyl Lewis a [50°], which is another ligand for E-selectin. The determination of the conformation bound to E-selectin has therefore been of particular interest. Figure 4 summarizes the conformations deduced from several transferred NOE studies [16",51,52]: all the studies predict that conformation A will be recognized by E-selectin, albeit with some variations in the local conformations. As the carbohydrate-binding domain of E-selectin has only been crystallized in it native state [53], there is no direct comparison available with a complexed form. Nevertheless, indirect structural data have been recently brought forward by the crystal structure of the complex of sLe x with the modified mannose-binding protein MBP-A [54°°]. This protein has been mutated to mimic the Ca2+-binding site of E-selectin and displays the same specificity and activity: In the complex, sLe x displays the same conformation as that predicted by the transferred N O E experiments [16°,51,52]. On the basis of the crystal structures of E-selectin [53] and MBP-oligosaccharide complexes [55], there have been several attempts to model the interaction between see x or its derivatives and selectins [56-58]. The agreement between these models and the selectin-like MBP/mutant model is poor, the source of the discrepancy being the orientation of the fucose in the primary binding site.
Oligosaccharide structures Imberty
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Comparison of (a) the MM3 energy maps of each disaccharide constituting sLe x together with (b) the conformations of the tetrasaccharide in different state. The shaded rectangle represents the sLe x conformations that are predicted to occur in a molecular mechanics study [47]. The four low-energy conformations are represented together with the nomenclature usually used [38]. The dots represent the conformation deduced from NMR study of the free tetrasaccharide [39,49]. The diamonds represent determined bound conformations in solution and are colored in black [51], gray [52] and white [16"]. The star indicates the conformation observed in the crystal complex with the selectin-like mutant of MBP-A [54*'].
Indeed, this exemplifies the difficulties and pitfalls in modeling protein-carbohydrate interactions.
References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as: • e.
Conclusions From the collective evidence I have reviewed, theoretical and experimental studies are clearly becoming mutually reliant for the elucidation of structural and dynamical data in the oligosaccharide field. On the one hand, even the most sophisticated N M R methods are insufficient to overcome the fact that experimental values represent only an average of the real solutions for flexible oligosaccharides [25•], and, thus, a thorough parallel modeling study is essential. On the other hand, most modeling research is now accompanied by complementary experimental validation, and it is this validation that contributes to the real value of the predicted model.
1.
Glennon TM, Zheng YJ, Le Grand SM, Shutzberg BA, Merz KM Jr: A force field for monosaccharides and (1-4) linked polysaccharides. J Comput Chem 1994, 15:1019-1040.
2.
Woods RJ, Dwek RA, Edge C J: Molecular mechanical and molecular dynamical simulations of glycoproteins and oligosaccharides. 1. GLYCAM-93 parameter developmenL J Am Chem Soc 1995, 99:3832-3846.
3.
Senderowitz H, Parish C, Still WC: Carbohydrates: united atom AMBER parametrization of pyranoses and simulations yielding anomeric free energies. J Am Chem Soc 1996, 118:2078-2086.
4.
Reiling S, Schlenkrich M, Brickrnann J: Force field parameters for carbohydrates. J Comput Chem 1996, 17:450-468.
5.
Ott KH, Meyer B: Parametrization of GROMOS force field for oligosaccharide assessment of efficiency of molecular dynamics simulations. J Comput Chem 1996, 17:1068-1084.
6.
Hardy BJ, Bystricky S, Kovac P, Wildmalm G: Conformational analysis and molecular dynamics simulation of c~-(1-2) and
Acknowledgements 1 gratefully acknowledge Serge P~rez for a fruitful scientific discussion, and Antoinette O'Sullivan for her careful reading of the manuscript.
of special interest of outstanding interest
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Carbohydrates and glycoconjugates
0c-(1-3) linked rhamnose oligosaccharides: reconciliation with optical rotation and NMR experiments. Biopo/ymers 1996, 41:83-96.
O-(0C-D-mannopyranosyl)-0C-D-mannopyranoside reveals two binding modes. J B/o/Chem 1996, 271:30614-30618. 19.
7. Ott KH, Meyer B: Molecular dynamics simulation of maltose in * water. CarbohydrRes 1996, 281:11-34. Extended MD simulations allow the determination of statistics on the conformational behavior of the o~GIc(1-4)GIc linkage. 8. Engelsen SB, P6rez S: The hydration of sucrose, Carbohydr Res •• 1996, 292:21-38. An analysis of a 500 ps MD simulation with water molecules which allows conclusions to be drawn concerning the hydration shell, the bridging water molecules, and their residence times. 9. •
Peters T, Pinto BM:Structure and dynamics of oligosaccharides: NMR and modeling studies. Curr Opin Struct Bio11996, 6:710-720. An in-depth review which includes oligosaccharides as part of glycoproteins and protein-carbohydrate complexes. Poveda A, Asensio JL, Espinosa JF, Martin-Pastor M, Canada J, Jimenez-Barbero J: Application of nuclear magnetic resonance spectroscopy and molecular modeling to the study of protein-carbohydrate interactions. J Mo/Graph 1997, 14:in press. This is an excellent review which covers all protein-carbohydrate interactions investigated by transferred NOEs.
20. =
Wright CS, Hester G: The 2.0/~ structure of a crosslinked complex between snowdrop lectin and a branched mannopentaose: evidence for two unique binding modes. Structure 1996 4:1339-1352. The conformation and binding mode of branched 3,6-core mannopentaose at two independent binding sites is described. Different conformations are observed primarily due to the flexibility about the ~ angle. 21.
Asensio JL, Jimenez-Barbero J: The use of the AMBER force field in conformational analysis of carbohydrate molecules: determination of the solution conformation of methyl c~-Iactoside by NMR spectroscopy, assisted by molecular mechanics and dynamics calculations. Biopo/ymers 1995, 35:55-73.
22.
Engelsen SB, Perez S, Braccini I, Herve du Penhoat C: Internal motions of carbohydrates as probed by comparative molecular modeling and nuclear magnetic resonance of ethyl ~-Iactoside. J Comput Chem 1995, 16:1096-1119.
10. .•
11. •
Casset F, Peters T, Etzler M, Korchagina E, Nifant'ev N, Perez S, Imberty A: Conformational analysis of blood group A trisaccharide in solution and in the binding site of Dolichos biflorus lectin using transient and transferred nuclear Overhauser enhancement (NOE) and rotating-frame NOE experiments. Eur J Biochem 1996, 239: 710-719. From a transferred-NOE investigation, lectin is concluded to select one conformation of blood group A trisaccharide from the equilibrium present in water solution. 12.
13.
Siebert HC, Gilleron M, Kaltner H, yon der Lieth CW, Kozar T, Bovin N, Korchagina EY, Vliegenthart .IF, Gabius HJ: NMR-based, molecular dynamics- and random walk molecular mechanicssupported study of conformational aspects of a carbohydrate ligand (Gal~l-2Gal ~I-R) for an animal galectin in the free and in the bound state. Biochem Biophys Res Comrnun 1996, 219:205-212. Espinosa JF, Canada FJ, Asensio JL, Dietrich H, Martin-Lomas M, Schmidt RR, Jimenes-Barbero J: Conformation differences of O and C-glycosides in the protein-bound state: different conformations of C-lactose and its O-analog are recognized by ricin B, a galactose-binding lectin. Angew Chem/nt Ed 1996, 35:303-306.
Espinosa -IF, Canada FJ, Asensio JL, Martin-Pastor M, Dietrich H, Martin-Lomas M, Schmidt RR, Jimenes-Barbero J: Experimental evidence of conformational differences between C-glycosides and O-glycosides in solution and in the protein-bound state: the C-lactose~O-lactose case. J Am Chem Soc 1996, 118:1 O862-10871, A transferred-NOEs investigation which concludes that ricin B selects different conformers of lactose and its analog, C-lactose. The flexibility of this C-disaccharide in solution is also demonstrated.
23. •
Martin-Pastor M, Espinosa .IF, Asensio JL, Jimenez-Barbero J: A comparison of the geometry and of the energy results obtained by application of different molecular mechanics force fields to methyl cx-lactoside and the C-analogue of lactose. Carbohydr Res 1997, 298:15-49. A comparison with clear conclusions about the force-fields that could be use for modeling carbohydrates. 24.
Casset F, Imberty A, Perez S, Etzler ME, Paulsen H, Peters T: Transferred nuclear Overhauser enhancement (NOE) and rotating-frame NOE experiments reflect the size of the bound segment of the Forssman pentasaccharide in the binding site of Dolichos biflorus lectin. Eur J Biochem 1997, 244:242-250. Ratio-dependent transferred NOE values are different for the buried and exposed extremities of the oligosaccharide. Within the binding site, the nature of the amino acid involved in each contact is determined by observation of ligand-protein transfer NOEs. 16. •
Poppe L, Brown GS, Philo JS, Nikrad PV, Shah BH: Conformation of sLex tetrasaccharide, free in solution and bound to E-, P-, and L-selectin. J Am Chem Soc 1997, 119:172-1736. The bound conformations of the ligand are calculated from transferred NOE data using the full relaxation matrix analysis, together with the ellipsoid model for the motion of the complex. The conformation of the ligand is predicted to be different when complexed with L-selectin than when complexed with E- and P-selectin. 17.
Naismith JH, Field IRA: Structural basis of trimannoside recognition by concanavalin A. J Biol Chem 1996, 271:972-976.
18.
Loris R, Maes D, Poortmans F, Wyns L, Bouckaert J: A structure of the complex between concanavalin A and methyl-3,6-di-
Espinosa JF, Martin-Pastor M, Asensio JL, Dietrich H, Martin-Lomas M, Schmidt RR, Jim*~nes-BarberoJ: Experimental and theoretical evidences of conformational flexibility of C-glycosides. NMR analysis and molecular mechanics calculations of C-lactose and its O-analogue. Tetrahedron Lett 1995, 36:6329-6332.
25. •
Rubinstenn G, Sinay P, Berthault P: Evidence of conformational heterogeneity for carbohydrate mimetics. NMR study of methyl 13-C-lactoside in aqueous solution. J Chem Phys A 1997, 101:2536-2540. The geometrical and dynamic parameters of 19 protons pairs are measured by off-resonance rotating-frame nuclear Overhauser enhancement spectroscopy. Direct determination of the conformation is not possible, however, because of structure flexibility. Combination of the NMR data with conformational analysis demonstrates the presence of two conformational families. 26.
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27.
Asensio JL, Canada FJ, Jimenez-Barbero J: Studies of the bound conformations of methyl oc-lactoside and methyl ~-gallactoside to ricin B chain using transferred NOE experiments in the laboratory and rotating frames, assisted by molecular mechanics and dynamics calculations. Eur J Biochem 1995, 233:618-630.
28.
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29.
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30.
Imberty A, Mikros E, Koca J, Mollicone R, Oriot R, Perez S: Computer simulation of histo-blood group oligosaccharides: energy maps of all constituting disaccharides and potential energy surfaces of 14 ABH and Lewis carbohydrate antigens. Glycoconj J 1995, 12:331-349.
31.
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32.
Longchambon F, Ohanessian J, Gillier-Pandraud H, Duchet D, Jacquinet -IC, Sinay P: Structure de la N-acetyl-lactosamine (ac6tamido-2-desoxy-2-O-~-D-galactopyrannosyl-4-c(-Dglucopyrannose). Acta Crystallogr B 1981, 37:601-607.
14. =•
15. •
Hester G, Wright CS: The mannose-specific bulb lectin from Galanfhus nivalis (snowdrop) binds mono- and dimannosides at distinct sites, Structure analysis of refined complexes at 2.3A and 3.0A resolution. J Mo/B/o/1996, 262:516-531.
Oligosaccharide structures Imberty
33.
Watt DK, Brasch DJ, Larsen DS, Melton LD, Simpson J: Oligosaccharides related to xyloglucan: synthesis and Xray crystal structure of methyl 2-O-(c(-I-fucopyranosyl)-~-Dgalactopyranoside. Carbohydr Res 1996, 285:1-15.
34.
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35.
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Duus JO, Nifant'ev N, Shashkov AS, Khatuntseva EA, Bock K: Synthesis and structural studies of branched 2-1inked trisaccharides related to blood group determinants. Carbohydr Res 1996, 288:25-44. Mollicone R, Cailleau A, Imberty A, Gane P, Perez S, Oriol R: Recognition of the blood group H type 2 trisaccharide epitope by 28 monoclonal antibodies and three lectins. Glycoconj J 1996, 13:263-271.
37. •
Imberty A, Mollicone R, Mikros E, Carrupt PA, Perez S, Oriol R: How do antibodies and lectins recognize histo-blood group antigens? A 3D-QSAR study by comparative molecular field analysis (CoMFA). Bioorg Med Chem 1996, 4:1979-1988, An extension of molecular modeling that demonstrates that quantitative structure/activity relationship (QSAR) methods can be used in the carbohydrate field. 38.
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39.
Miller KE, Mukhopadhyay C, Cagas P, Bush CA: Solution structure of the Lewis x oligosaccharide determined by N M R spectroscopy and molecular dynamics simulations. Biochemistry 1992, 31:6703-6709.
40.
Poveda A, Asensio JL, Martin-Pastor M, Jimenez-Barbero J: Exploration of the conformational flexibility of the Lex related oligosaccharide GalNAcc¢(1-3) Gall~(1-4)[Fuccx(1-3)]GIc by 1H NMR relaxation measurements and molecular dynamics simulations. Chem Comrnun 1996, 421-422.
41. •
Poveda A, Asensio JL, Martin-Pastor M, Jimenez-Barbero J: Solution conformation and dynamics of a tetrasaccharide related to the Lewis x antigen deduced by NMR relaxation measurements. J Biomol NMR 1997, in press. 1H and 13C relaxation data obtained at two different fields and a variety of temperatures permit the determination of local dynamical parameters. A comparison of these with MD simulation yields a fair agreement. 42.
Berthault P, Birtikaris N, Rubinstenn G, Sinay P, Desvaux H: Solution structure of a Lewis x analogue by off-resonance 1H N M R spectroscopy without use of an internal distance reference. J Biomol NMR 1996, 8:23-35. The use of off-resonance rotative frame nuclear Overhauser enhancement spectroscopy allows the determination of geometrical and dynamical parameters of 30 proton pairs.
47.
Imberty A, Perez S: Traveling through the potential energy surface of sialyl Lewis x. In Carbohydrate Mimics. Concepts and Methods. Edited by Chapleur Y. Weinheim: Wiley; 1997:349-364.
48.
Mukhopadhyay C, Miller KE, Bush CA:Conformation of the oligosaccharide receptor for E-selectin. Biopo/ymers 1994, 34:21-29.
49.
Rutherford TJ, Spackman DG, Simpson PJ, Homans SW: 5 nanosecond molecular dynamics and NMR study of conformational transitions in the sialyI-Lewis x antigen. G/ycobio/ogy 1994, 4:59-68.
50. •
Kogelberg H, Frenkiel TA, Homans SW, Lubineau A, Feizi T: Conformational studies on the selectin and natural killer cell receptor ligands sulfo- and sialyl-lacto-N-fucopentaoses (SuLNFPII and SLNFPII) using NMR spectroscopy and molecular dynamics simulations. Comparisons with the nonacidic parent molecule LNFPII. Biochemistry 1996, 35:1954-1 g64. Rotative-frame nuclear Overhauser enhancement spectroscopy and long-range 1H,13C-coupling constants are used together with MD simulation to determine the conformational behavior of several Lea-containing oligosaccharides. 51.
Cooke RM, Hale RS, Lister SG, Shah G, Weir MP: The conformation of the sialyl Lewis x ligand changes upon binding to E-selectin. Biochemistry 1 g94, 33:1 O59-10596.
52.
Scheffler K, Ernst B, Katopodis A, Magnani JL, Wang WT, Weisemann R, Peters T: Determination of the carbohydrate ligand in the E-selectin-sialyl Lewis x complex. Angew Chem Int Ed 1995, 34:184-1844.
53.
Graves BJ, Crowther RL, Chandran C, Rumberger JM, Li S, Huang KS, Presky DH, Familletti PC, Wolitzky BA, Burns DK: Insight into E-selectin-ligand interaction from the crystal structure and mutagenesis of the lec/EGF domains. Nature 1994, 367:532-538.
54. ••
Ng KK-S, Weis Wl: Structure of a selectin-like mutant of mannose-binding protein complexed with sialylated and sulfated Lewis x oligosaccharides. Biochemistry 1997, 36:979-988. An indirect structural approach to investigate the interaction between oligosaccharide and E-selectin. The conformation of sLex in the crystalline complex is in good agreement with the bound conformation in solution [16°,51,52]. The lack of observed interaction between NeuNAc and the protein is surprising and opens a wide range of issues. 55.
Weis Wl, Drickamer K, Hendrickson WA:Structure of a C-type mannose-binding protein complexed with an oligosaccharide. Nature 1992, 360:127-134.
56.
Kogan TP, Revelle BM, Tapp S, Scott D, Beck PJ: A single amino acid residue can determine the ligand specificity of E-selectin. J Biol Chem 1995, 270:14047-14055.
57.
Hiramatsu Y, Tsujishita H, Kondo H: Studies on selectin blockers. 3. Investigation of the carbohydrate ligand sialyl Lewis x recognition site of P-selectin. J Med Chem 1996, 39:4547-4553.
58.
Tsujishita H, Hiramatsu Y, Kondo N, Ohmoto H, Kondo H, Kiso M, Hasegawa A: Selectin-ligand interactions revealed by molecular dynamics simulation in solution. J Med Chem 1997, 40:362-369.
59.
Sokolowski T, Peters T, Perez S, Imberty A: Conformational analysis of biantennary glycans and molecular modeling of their complexes with lentil lectin. J Mo/Graph 1997, 15:in press.
60.
Bourne Y, Mazurier J, Legrand D, Rouge P, Montreuil J, Spik G, Cambillau C: Structures of a legume lectin complexed with the human lactotransferrin N2 fragment, and with an isolated biantennary glycopeptide: role of the fucose moiety. Structure 1994, 2:209-219.
61.
Bourne Y, Rough P, Cambillau C: X-ray structure of a biantennary octasaccharide-lectin complex refined at 2.3/~, resolution. J Biol Chem 1992, 267:19?-203.
62.
Bourne Y, Bolgiano B, Liao DI, Strecker G, Cantau P, Herzberg O, Feizi T, Cambillau C: Crosslinking of mammalian lectin (galectin-1) by complex biantennary saccharides. Nat Struct Biol 1994, 1:863-870.
•
43. ••
Perez S, Mouhous-Riou N, Nifant'ev NE, Tsvetkov YE, Bachet B, Imberty A:Crystal and molecular structure of a histoblood group antigen involved in cell adhesion: the Lewis x trisaccharide. Glycobiology 1996, 6:537~542. The first description of the crystal structure of a histo-blood group carbohydrate determinant. Some interactions observed between adjacent trisaccharides can provide clues for carbohydrate-carbohydrate interactions in cell adhesion. 44.
YvelinF, Zhang YM, Mallet JM, Robert F, Jeannin Y, Sinay P: Crystal structure of the Lewis x trisaccharide. Carbohydr Lett 1996, 1:475-482.
45.
Kojima N, Fenderson BA, Stroud MR, Goldberg RI, Habermann R, Toyokuni T, Hakomori S: Further studies on cell adhesion based on LeX-Lex interaction, with new approaches: embryoglycan aggregation of F9 teratocarcinoma cells, and adhesion of various turnout cells based on Lex expression. G/ycoconj -I 1994, 11:238-248.
46.
Ni F: Complete relaxation analysis matrix of transferred nuclear Overhauser effects. J Magn Reson 1 992, 96:651-656.
623
Oligosaccharide structures: theory versus experiment Anne Imberty Recently, the interdependency of theoretical and experimental approaches in the structure determination of oligosaccharides has been confirmed. More accurate simulations are possible because of the advances in software and computers. Meanwhile, improvements in NMR techniques permit the measurement of numerous structural and dynamical parameters, either for the free state or for carbohydrate ligands bound to receptors. Several crystal structures of isolated or protein-complexed oligosaccharides give new clues for modeling the intermolecular forces that drive the interactions.
Addresses Centre de Recherches sur les Macromolacules Veg6tales, Joseph Fourier Universite, BP53, F38041, Grenoble cedex 9, France; e-mail: [email protected] Current Opinion in Structural Biology 1997, 7:61 ?-623 http://biomednet.com/elecref/O959440X0070061 ? © Current Biology Ltd ISSN 0959-440X Abbreviations MBP mannose-binding protein MD molecular dynamics NOE nuclear Overhauser effect sLe x sialyl Lewisx
Introduction In many instances, even extensive knowledge of the primary structure of oligosaccharides is insufficient to understand and explain their function and specificity. T h e 3D structures of glycans provide the driving force for all intermolecular interactions and therefore predetermine their function; furthermore, the flexibility and dynamics of oligosaccharides frequently play a key role in biological activity and must also be taken into account. In recent years, it has been recognized that efficient carbohydrate modeling demands the use of appropriate force-fields, as conformational preference at the glycosidic linkage is due to special stereoelectronic effects. Some of the general force-fields, such as MM3, have proven to be suitable for this purpose, whereas others require supplementary parametrization. During the past two years, several new parametrizations for carbohydrates have been proposed for the programs more widely used in the domain of molecular dynamics (MD). Examples can be found for AMBER [1-3], C H A R M m [41 and GROMOS [5]. As a result of theses advances, some extended MD simulations have been routinely calculated for disaccharides [6], and the inclusion of explicit water molecules has been facilitated [7",8°°].
Even if modeling groups are generally aware of the conformational particularities of carbohydrates, one must keep in mind the fact that any model proposed via a theoretical study should, where possible, be validated by a comparison with experimental data. X-ray crystallography provides the most precise information on the shape of a carbohydrate molecule. In addition, a thorough analysis of packing arrangements frequently produces information on preferential carbohydrate-carbohydrate and carbohydrate-water relationships. A limitation of the above method is that it does not reflect oligosaccharide flexibility. A more fundamental point to be considered is the difficulty encountered in the crystallization of oligosaccharides and even disaccharides. N M R experiments are widely used for the validation of structures of oligosaccharides using predicted molecular modeling. Measurements of nuclear Overhauser effects (NOEs) help in the determination of conformations in solution and, in addition, have the tremendous advantage of giving access to some dynamic properties of molecules such as the timescales of internal motions. During the past two years, the major advances have been in the determination of the bioactive conformation of oligosaccharides, that is, their conformation when bound to a protein receptor. In solution, transferred N O E experiments have proven to be a powerful tool in the investigation of the bound conformations of oligosaccharides [9",10°°]. In the past year, this method has been successfully used as a tool for the conformational analysis of bound carbohydrates and analogs [I 1",12,13,14°°,15°,16°]. In the mean time, several new crystal structures of plant lectin-oligosaccharide complexes have been solved [17-19,20°]. For some linkages, such as those of the Manc~(1-3)Man and GIcNAc[3(1-2)Man disaccharides, crystal structures of lectin-oligosaccharide complexes provide sufficient evidence to illustrate that the conformations selected upon binding indeed correspond to the several energy minima predicted by theoretical calculations performed on individual glycosidic linkages (Figure 1). I will review several recent examples of oligosaccharide structure for which both theoretical and experimental data are available.
Comparison of the conformation of lactose and its C-analog T h e conformational behavior of the lactose disaccharide (]3Gal[1-4]GIc) has been well characterized using a combination of theoretical and experimental methods. Its potential energy surface is characteristic of equatorial-equatorial (1-4)-linked disaccharides possessing three or four major potential-energy wells. T h e most populated conformation
618
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is the so-called syn, which corresponds to all observed conformations of lactose crystal structures, including those found in protein complexes. Anti and gauche-gauche conformations differ from the lowest energy shape by a large change in the ~) and gt torsion angles, respectively [21,22]. As each of the three conformations has a characteristic NOE fingerprint (Figure 2), it has been possible to deduce that the population of lactose in solution is about 90% svn and 10% anti, with no detectable gauche-gauche conformer [14"*,21]. Because of its well-characterized conformational behavior, lactose is now used for testing force-fields [22,23"]. Recentl'> C-disaccharides have been investigated as potential mimetics of natural disaccharides. As the glycosidic oxygen is replaced by a methylene group, these compounds exhibit a higher chemical and biochemical stability. A significant interest has arisen in comparing the molecular mechanics and dynamics behavior of C-disaccharide with those of O-disaccharide to determine whether such compounds can act as competitive inhibitors of their O-counterparts. A conformational study of C-lactose using a combination of NMR spectroscopy and molecular mechanics has shown that it is more flexible than lactose [24]. Three main low-energy conformations ate predicted to coexist in solution with a distribution of 55%, 40%, and 5% for syn, anti, and gauche-gauche, respectively. Another study has corroborated the flexibility of C-lactose [25"],
but the comparison of experimental and back-calculated N O E values predicts the presence of only two conformers, namely syn (60%) and anti (40%). Both O-lactose and C-lactose conformations have also been studied in the protein-bound state. When methyl [3-1actoside [26] or methyl o~-lactoside [27] are bound to ricin B, the measured transferred N O E implies only a slight variation from the lowest energy anti conformation. T h e situation is completely different with C-lactose. In the presence of ricin B, only the transferred NOE characteristic of the syn conformation can be observed [13,14"]. These experimental results indicate, therefore, that ricin B selects different conformers of C-lactose and O-lactose. Thus, it appears that it is possible for the inhibitor conformation selected by the protein receptor to be different from that of the natural substrate. Such conformational behavior could prove to be vital when designing new oligosaccharide mimics. Histo-blood group oligosaccharides Conformational analysis of histo-blood group carbohydrate determinants was pioneered by Lemieux et al. [28] and Rao and Biswas [29] more than 15 years ago (Figure 3). These molecules have been the subject of much interest in recent years, while the role of the Lewis determinants has been discovered in many biological functions. Conformational analysis of all constituting disaccharides
Oligosaccharide structures Imberty
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of blood group oligosaccharides has been performed recently [30,31]. For comparison, the only disaccharide fragments for which crystal structures are available arc N-acetyllactosamine [32], which was crystallized more than 10 years ago, and the H-type disaccharide otFuc(1-2)13Gal (blood group O determinant) [33]. In both cases, the crystal conformation corresponds to the predicted lowest energy regions. All the constituent disaccharides are predicted to be flexible. T h e conformational behavior of histo-blood group oligosaccharides is different. Depending on their branch-
ing patterns, they can exhibit a range of conformational behavior, from almost complete rigidity to high flexibility [30,34,35]. T h e different alignments of oligosaccharides that have been deduced from these studies have been used to determine the epitopes recognized by monoclonal antibodies [36]. In a more in-depth study, 3D quantitative structure/activity relationship (QSAR) methods have been applied to these molecules in order to understand the cross-reaction patterns of antiblood group antibodies [37°]. In this case, the agreement of molecular mechanics models with immunological data represents an experimental validation of the theoretical study.
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620
Carbohydrates and glycoconjugates
The Lewis x trisaccharide
The Lewis x determinant, [3Gal(1-4)lotFuc(1-3)]13GlcNAc, is a trisaccharidc of biological interest as it is a stagespecific embryonic marker involved in cell-cell interactions and a signaling molecule for some host-pathogen recognition. All the theoretical studies [28-31] have predicted one principal low-energy conformation, characterized by a stacking between the fucosc and galactose rings. The otFuc(1-3)13GlcNAc linkage conformation corresponds to one of the two lowest energy minima for the disaccharide, whereas the fucose-galactose interaction shifts the I3Gal(1-4)I3GlcNAc linkage to the border of a conformational plateau. Other conformational families only possibly exist at significantly higher energies; however, they can occur in special cases, such as interactions with a receptor. The conformation predicted from theoretical studies has been fully confirmed by NMR experiments on the Lewis x trisaccharide [38,39].
otGalNAc(1-3)[otFuc(1-2)]13Gal, two main conformations are predicted to occur, which differ mainly by the orientation of the fucose residue. Comparison of the simulated build-up NOE curves with experimental ones [11"] permits the confirmation of the presence of an equilibrium between these two conformers. Interaction of blood group A determinant with the lcctin from Dolichos biflorus seeds has been characterized by negative transferred NOEs corresponding only to one conformation of the trisaccharide. Theoretical transferred NOEs, calculated using an extended procedure for the complete relaxation matrix analysis of multispin exchanging systems [46], have confirmed that only one conformation is selected for binding the lectin. This result is not obvious from molecular modeling alone, as both solution conformations can be satisfactorily docked in the binding site of the D. biflorus lectin. Sialyl Lewis x
Interestingly, NMR studies performed on analogs have demonstrated a larger flexibility. A thorough 1H NMR relaxation study coupled with MD simulation [40,41 °] performed on otGalNAc(1-3)[3Gal-(1-4)[o~Fuc(1-3)]13GlcOMe has indicated that the fucosc is more mobile, presumably because of the lack of an acetamido group on the reducing glucose. The same conclusions have been reached for an analog in which the O-glycosidic bond linking fucose to glucose has been replaced by a C-glycosidic bond [42"]. In this case, longitudinal and transverse dipolar cross-relaxation rates have been measured with great accuracy, allowing a direct derivation of both geometrical and dynamical parameters. From a conformational point of vie~, therefore, studying the crystal structure of such a rigid compound may seem pointless. Nevertheless, some very interesting points have arisen from the resolution of the crystal structure [43"',44]. First, the comparison of the two independent molecules contained in the asymmetric unit has indicated that both adopt the global low-energy conformation, but with variations of - 1 0 ° in the torsion angles of the glycosidic linkages thereby confirming the local flexibility predicted by modeling and NMR studies. Second, the structure contains nine water molecules in the asymmetric unit, which is very unusual in a carbohydrate crystal structure. Lastly, the interactions between adjacent trisaccharides that are observed in the crystal packing correspond to LcX-Le x interactions that could bc involved in the formations of glycosidic clusters at the surfaces of cells and in carbohydrate-mediated cell-cell interactions [45]. The crystal structure is, therefore, a possible template for further modeling of carbohydrate-carbohydrate interactions. The blood group A determinant
Initially described as a rigid molecule [28,29], blood group A determinant has now been proposed to have several possible conformations, as predicted by a molecular mechanics study [30]. For the blood group A trisaccharide
Sialyl Lewis x (sLe x) has been the subject of intensive structural studies during the past few years because of its involvement in cellular adhesion. Molecular modeling studies agree in regard to the description of the tetrasaccharide being a rigid core consisting of the Lewis x moiety attached to sialic acid via a rather flexible linkage--two or three conformations being accessible to the molecule [34,38,47]. NMR studies with NOE measurements of the free tetrasaccharide all agree on the flexibility of the sialic acid moiety [16°,48,49]. The same conclusion has been derived from a NMR spectroscopy and molecular dynamics study of sialyl Lewis a [50°], which is another ligand for E-selectin. The determination of the conformation bound to E-selectin has therefore been of particular interest. Figure 4 summarizes the conformations deduced from several transferred NOE studies [16",51,52]: all the studies predict that conformation A will be recognized by E-selectin, albeit with some variations in the local conformations. As the carbohydrate-binding domain of E-selectin has only been crystallized in it native state [53], there is no direct comparison available with a complexed form. Nevertheless, indirect structural data have been recently brought forward by the crystal structure of the complex of sLe x with the modified mannose-binding protein MBP-A [54°°]. This protein has been mutated to mimic the Ca2+-binding site of E-selectin and displays the same specificity and activity: In the complex, sLe x displays the same conformation as that predicted by the transferred N O E experiments [16°,51,52]. On the basis of the crystal structures of E-selectin [53] and MBP-oligosaccharide complexes [55], there have been several attempts to model the interaction between see x or its derivatives and selectins [56-58]. The agreement between these models and the selectin-like MBP/mutant model is poor, the source of the discrepancy being the orientation of the fucose in the primary binding site.
Oligosaccharide structures Imberty
621
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Comparison of (a) the MM3 energy maps of each disaccharide constituting sLe x together with (b) the conformations of the tetrasaccharide in different state. The shaded rectangle represents the sLe x conformations that are predicted to occur in a molecular mechanics study [47]. The four low-energy conformations are represented together with the nomenclature usually used [38]. The dots represent the conformation deduced from NMR study of the free tetrasaccharide [39,49]. The diamonds represent determined bound conformations in solution and are colored in black [51], gray [52] and white [16"]. The star indicates the conformation observed in the crystal complex with the selectin-like mutant of MBP-A [54*'].
Indeed, this exemplifies the difficulties and pitfalls in modeling protein-carbohydrate interactions.
References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as: • e.
Conclusions From the collective evidence I have reviewed, theoretical and experimental studies are clearly becoming mutually reliant for the elucidation of structural and dynamical data in the oligosaccharide field. On the one hand, even the most sophisticated N M R methods are insufficient to overcome the fact that experimental values represent only an average of the real solutions for flexible oligosaccharides [25•], and, thus, a thorough parallel modeling study is essential. On the other hand, most modeling research is now accompanied by complementary experimental validation, and it is this validation that contributes to the real value of the predicted model.
1.
Glennon TM, Zheng YJ, Le Grand SM, Shutzberg BA, Merz KM Jr: A force field for monosaccharides and (1-4) linked polysaccharides. J Comput Chem 1994, 15:1019-1040.
2.
Woods RJ, Dwek RA, Edge C J: Molecular mechanical and molecular dynamical simulations of glycoproteins and oligosaccharides. 1. GLYCAM-93 parameter developmenL J Am Chem Soc 1995, 99:3832-3846.
3.
Senderowitz H, Parish C, Still WC: Carbohydrates: united atom AMBER parametrization of pyranoses and simulations yielding anomeric free energies. J Am Chem Soc 1996, 118:2078-2086.
4.
Reiling S, Schlenkrich M, Brickrnann J: Force field parameters for carbohydrates. J Comput Chem 1996, 17:450-468.
5.
Ott KH, Meyer B: Parametrization of GROMOS force field for oligosaccharide assessment of efficiency of molecular dynamics simulations. J Comput Chem 1996, 17:1068-1084.
6.
Hardy BJ, Bystricky S, Kovac P, Wildmalm G: Conformational analysis and molecular dynamics simulation of c~-(1-2) and
Acknowledgements 1 gratefully acknowledge Serge P~rez for a fruitful scientific discussion, and Antoinette O'Sullivan for her careful reading of the manuscript.
of special interest of outstanding interest
622
Carbohydrates and glycoconjugates
0c-(1-3) linked rhamnose oligosaccharides: reconciliation with optical rotation and NMR experiments. Biopo/ymers 1996, 41:83-96.
O-(0C-D-mannopyranosyl)-0C-D-mannopyranoside reveals two binding modes. J B/o/Chem 1996, 271:30614-30618. 19.
7. Ott KH, Meyer B: Molecular dynamics simulation of maltose in * water. CarbohydrRes 1996, 281:11-34. Extended MD simulations allow the determination of statistics on the conformational behavior of the o~GIc(1-4)GIc linkage. 8. Engelsen SB, P6rez S: The hydration of sucrose, Carbohydr Res •• 1996, 292:21-38. An analysis of a 500 ps MD simulation with water molecules which allows conclusions to be drawn concerning the hydration shell, the bridging water molecules, and their residence times. 9. •
Peters T, Pinto BM:Structure and dynamics of oligosaccharides: NMR and modeling studies. Curr Opin Struct Bio11996, 6:710-720. An in-depth review which includes oligosaccharides as part of glycoproteins and protein-carbohydrate complexes. Poveda A, Asensio JL, Espinosa JF, Martin-Pastor M, Canada J, Jimenez-Barbero J: Application of nuclear magnetic resonance spectroscopy and molecular modeling to the study of protein-carbohydrate interactions. J Mo/Graph 1997, 14:in press. This is an excellent review which covers all protein-carbohydrate interactions investigated by transferred NOEs.
20. =
Wright CS, Hester G: The 2.0/~ structure of a crosslinked complex between snowdrop lectin and a branched mannopentaose: evidence for two unique binding modes. Structure 1996 4:1339-1352. The conformation and binding mode of branched 3,6-core mannopentaose at two independent binding sites is described. Different conformations are observed primarily due to the flexibility about the ~ angle. 21.
Asensio JL, Jimenez-Barbero J: The use of the AMBER force field in conformational analysis of carbohydrate molecules: determination of the solution conformation of methyl c~-Iactoside by NMR spectroscopy, assisted by molecular mechanics and dynamics calculations. Biopo/ymers 1995, 35:55-73.
22.
Engelsen SB, Perez S, Braccini I, Herve du Penhoat C: Internal motions of carbohydrates as probed by comparative molecular modeling and nuclear magnetic resonance of ethyl ~-Iactoside. J Comput Chem 1995, 16:1096-1119.
10. .•
11. •
Casset F, Peters T, Etzler M, Korchagina E, Nifant'ev N, Perez S, Imberty A: Conformational analysis of blood group A trisaccharide in solution and in the binding site of Dolichos biflorus lectin using transient and transferred nuclear Overhauser enhancement (NOE) and rotating-frame NOE experiments. Eur J Biochem 1996, 239: 710-719. From a transferred-NOE investigation, lectin is concluded to select one conformation of blood group A trisaccharide from the equilibrium present in water solution. 12.
13.
Siebert HC, Gilleron M, Kaltner H, yon der Lieth CW, Kozar T, Bovin N, Korchagina EY, Vliegenthart .IF, Gabius HJ: NMR-based, molecular dynamics- and random walk molecular mechanicssupported study of conformational aspects of a carbohydrate ligand (Gal~l-2Gal ~I-R) for an animal galectin in the free and in the bound state. Biochem Biophys Res Comrnun 1996, 219:205-212. Espinosa JF, Canada FJ, Asensio JL, Dietrich H, Martin-Lomas M, Schmidt RR, Jimenes-Barbero J: Conformation differences of O and C-glycosides in the protein-bound state: different conformations of C-lactose and its O-analog are recognized by ricin B, a galactose-binding lectin. Angew Chem/nt Ed 1996, 35:303-306.
Espinosa -IF, Canada FJ, Asensio JL, Martin-Pastor M, Dietrich H, Martin-Lomas M, Schmidt RR, Jimenes-Barbero J: Experimental evidence of conformational differences between C-glycosides and O-glycosides in solution and in the protein-bound state: the C-lactose~O-lactose case. J Am Chem Soc 1996, 118:1 O862-10871, A transferred-NOEs investigation which concludes that ricin B selects different conformers of lactose and its analog, C-lactose. The flexibility of this C-disaccharide in solution is also demonstrated.
23. •
Martin-Pastor M, Espinosa .IF, Asensio JL, Jimenez-Barbero J: A comparison of the geometry and of the energy results obtained by application of different molecular mechanics force fields to methyl cx-lactoside and the C-analogue of lactose. Carbohydr Res 1997, 298:15-49. A comparison with clear conclusions about the force-fields that could be use for modeling carbohydrates. 24.
Casset F, Imberty A, Perez S, Etzler ME, Paulsen H, Peters T: Transferred nuclear Overhauser enhancement (NOE) and rotating-frame NOE experiments reflect the size of the bound segment of the Forssman pentasaccharide in the binding site of Dolichos biflorus lectin. Eur J Biochem 1997, 244:242-250. Ratio-dependent transferred NOE values are different for the buried and exposed extremities of the oligosaccharide. Within the binding site, the nature of the amino acid involved in each contact is determined by observation of ligand-protein transfer NOEs. 16. •
Poppe L, Brown GS, Philo JS, Nikrad PV, Shah BH: Conformation of sLex tetrasaccharide, free in solution and bound to E-, P-, and L-selectin. J Am Chem Soc 1997, 119:172-1736. The bound conformations of the ligand are calculated from transferred NOE data using the full relaxation matrix analysis, together with the ellipsoid model for the motion of the complex. The conformation of the ligand is predicted to be different when complexed with L-selectin than when complexed with E- and P-selectin. 17.
Naismith JH, Field IRA: Structural basis of trimannoside recognition by concanavalin A. J Biol Chem 1996, 271:972-976.
18.
Loris R, Maes D, Poortmans F, Wyns L, Bouckaert J: A structure of the complex between concanavalin A and methyl-3,6-di-
Espinosa JF, Martin-Pastor M, Asensio JL, Dietrich H, Martin-Lomas M, Schmidt RR, Jim*~nes-BarberoJ: Experimental and theoretical evidences of conformational flexibility of C-glycosides. NMR analysis and molecular mechanics calculations of C-lactose and its O-analogue. Tetrahedron Lett 1995, 36:6329-6332.
25. •
Rubinstenn G, Sinay P, Berthault P: Evidence of conformational heterogeneity for carbohydrate mimetics. NMR study of methyl 13-C-lactoside in aqueous solution. J Chem Phys A 1997, 101:2536-2540. The geometrical and dynamic parameters of 19 protons pairs are measured by off-resonance rotating-frame nuclear Overhauser enhancement spectroscopy. Direct determination of the conformation is not possible, however, because of structure flexibility. Combination of the NMR data with conformational analysis demonstrates the presence of two conformational families. 26.
Bevilacqua VL, Thomson DS, Prestegard JH: Conformation of methyl ~-Iactoside bound to the ricin B-chain: interpretation of transferred nuclear Overhauser effects facilitated by spin simulation and selective deutaration. Biochemistry 1990, 29:5529-5537
27.
Asensio JL, Canada FJ, Jimenez-Barbero J: Studies of the bound conformations of methyl oc-lactoside and methyl ~-gallactoside to ricin B chain using transferred NOE experiments in the laboratory and rotating frames, assisted by molecular mechanics and dynamics calculations. Eur J Biochem 1995, 233:618-630.
28.
Lemieux RU, Bock K, Delbaere LTJ, Koto S, Rao VS: The conformations of oligosaccharides related to the ABH and Lewis human blood group determinants. Can J Chem 1980, 58:63-65.
29.
Rao VSR, Biswas M: Conformations and interactions of oligosaccharides related to the ABH and Lewis blood groups. Top Mol Struct Biol 1985, 8:185-218.
30.
Imberty A, Mikros E, Koca J, Mollicone R, Oriot R, Perez S: Computer simulation of histo-blood group oligosaccharides: energy maps of all constituting disaccharides and potential energy surfaces of 14 ABH and Lewis carbohydrate antigens. Glycoconj J 1995, 12:331-349.
31.
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14. =•
15. •
Hester G, Wright CS: The mannose-specific bulb lectin from Galanfhus nivalis (snowdrop) binds mono- and dimannosides at distinct sites, Structure analysis of refined complexes at 2.3A and 3.0A resolution. J Mo/B/o/1996, 262:516-531.
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33.
Watt DK, Brasch DJ, Larsen DS, Melton LD, Simpson J: Oligosaccharides related to xyloglucan: synthesis and Xray crystal structure of methyl 2-O-(c(-I-fucopyranosyl)-~-Dgalactopyranoside. Carbohydr Res 1996, 285:1-15.
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Bizik F, Tvaroska I: On the flexibility of the Lewis x, Lewis a, sialyl Lewis x and sialyl Lewisa oligosaccharides. Conformational analysis in solution by molecular modelling. Chem Papers 1996, 50:84-86.
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Duus JO, Nifant'ev N, Shashkov AS, Khatuntseva EA, Bock K: Synthesis and structural studies of branched 2-1inked trisaccharides related to blood group determinants. Carbohydr Res 1996, 288:25-44. Mollicone R, Cailleau A, Imberty A, Gane P, Perez S, Oriol R: Recognition of the blood group H type 2 trisaccharide epitope by 28 monoclonal antibodies and three lectins. Glycoconj J 1996, 13:263-271.
37. •
Imberty A, Mollicone R, Mikros E, Carrupt PA, Perez S, Oriol R: How do antibodies and lectins recognize histo-blood group antigens? A 3D-QSAR study by comparative molecular field analysis (CoMFA). Bioorg Med Chem 1996, 4:1979-1988, An extension of molecular modeling that demonstrates that quantitative structure/activity relationship (QSAR) methods can be used in the carbohydrate field. 38.
Ichikawa K, Lin YC, Dumas DP, Shen G J, Garcia-Junceda E, Williams MA, Bayer R, Ketcham C, Walker LE, Paulson JC, Wonc CH: Chemical-enzymatic synthesis and conformational analysis of sialyl Lewis x and derivatives. J Am Chem Soc 1992, 114:9283-9288.
39.
Miller KE, Mukhopadhyay C, Cagas P, Bush CA: Solution structure of the Lewis x oligosaccharide determined by N M R spectroscopy and molecular dynamics simulations. Biochemistry 1992, 31:6703-6709.
40.
Poveda A, Asensio JL, Martin-Pastor M, Jimenez-Barbero J: Exploration of the conformational flexibility of the Lex related oligosaccharide GalNAcc¢(1-3) Gall~(1-4)[Fuccx(1-3)]GIc by 1H NMR relaxation measurements and molecular dynamics simulations. Chem Comrnun 1996, 421-422.
41. •
Poveda A, Asensio JL, Martin-Pastor M, Jimenez-Barbero J: Solution conformation and dynamics of a tetrasaccharide related to the Lewis x antigen deduced by NMR relaxation measurements. J Biomol NMR 1997, in press. 1H and 13C relaxation data obtained at two different fields and a variety of temperatures permit the determination of local dynamical parameters. A comparison of these with MD simulation yields a fair agreement. 42.
Berthault P, Birtikaris N, Rubinstenn G, Sinay P, Desvaux H: Solution structure of a Lewis x analogue by off-resonance 1H N M R spectroscopy without use of an internal distance reference. J Biomol NMR 1996, 8:23-35. The use of off-resonance rotative frame nuclear Overhauser enhancement spectroscopy allows the determination of geometrical and dynamical parameters of 30 proton pairs.
47.
Imberty A, Perez S: Traveling through the potential energy surface of sialyl Lewis x. In Carbohydrate Mimics. Concepts and Methods. Edited by Chapleur Y. Weinheim: Wiley; 1997:349-364.
48.
Mukhopadhyay C, Miller KE, Bush CA:Conformation of the oligosaccharide receptor for E-selectin. Biopo/ymers 1994, 34:21-29.
49.
Rutherford TJ, Spackman DG, Simpson PJ, Homans SW: 5 nanosecond molecular dynamics and NMR study of conformational transitions in the sialyI-Lewis x antigen. G/ycobio/ogy 1994, 4:59-68.
50. •
Kogelberg H, Frenkiel TA, Homans SW, Lubineau A, Feizi T: Conformational studies on the selectin and natural killer cell receptor ligands sulfo- and sialyl-lacto-N-fucopentaoses (SuLNFPII and SLNFPII) using NMR spectroscopy and molecular dynamics simulations. Comparisons with the nonacidic parent molecule LNFPII. Biochemistry 1996, 35:1954-1 g64. Rotative-frame nuclear Overhauser enhancement spectroscopy and long-range 1H,13C-coupling constants are used together with MD simulation to determine the conformational behavior of several Lea-containing oligosaccharides. 51.
Cooke RM, Hale RS, Lister SG, Shah G, Weir MP: The conformation of the sialyl Lewis x ligand changes upon binding to E-selectin. Biochemistry 1 g94, 33:1 O59-10596.
52.
Scheffler K, Ernst B, Katopodis A, Magnani JL, Wang WT, Weisemann R, Peters T: Determination of the carbohydrate ligand in the E-selectin-sialyl Lewis x complex. Angew Chem Int Ed 1995, 34:184-1844.
53.
Graves BJ, Crowther RL, Chandran C, Rumberger JM, Li S, Huang KS, Presky DH, Familletti PC, Wolitzky BA, Burns DK: Insight into E-selectin-ligand interaction from the crystal structure and mutagenesis of the lec/EGF domains. Nature 1994, 367:532-538.
54. ••
Ng KK-S, Weis Wl: Structure of a selectin-like mutant of mannose-binding protein complexed with sialylated and sulfated Lewis x oligosaccharides. Biochemistry 1997, 36:979-988. An indirect structural approach to investigate the interaction between oligosaccharide and E-selectin. The conformation of sLex in the crystalline complex is in good agreement with the bound conformation in solution [16°,51,52]. The lack of observed interaction between NeuNAc and the protein is surprising and opens a wide range of issues. 55.
Weis Wl, Drickamer K, Hendrickson WA:Structure of a C-type mannose-binding protein complexed with an oligosaccharide. Nature 1992, 360:127-134.
56.
Kogan TP, Revelle BM, Tapp S, Scott D, Beck PJ: A single amino acid residue can determine the ligand specificity of E-selectin. J Biol Chem 1995, 270:14047-14055.
57.
Hiramatsu Y, Tsujishita H, Kondo H: Studies on selectin blockers. 3. Investigation of the carbohydrate ligand sialyl Lewis x recognition site of P-selectin. J Med Chem 1996, 39:4547-4553.
58.
Tsujishita H, Hiramatsu Y, Kondo N, Ohmoto H, Kondo H, Kiso M, Hasegawa A: Selectin-ligand interactions revealed by molecular dynamics simulation in solution. J Med Chem 1997, 40:362-369.
59.
Sokolowski T, Peters T, Perez S, Imberty A: Conformational analysis of biantennary glycans and molecular modeling of their complexes with lentil lectin. J Mo/Graph 1997, 15:in press.
60.
Bourne Y, Mazurier J, Legrand D, Rouge P, Montreuil J, Spik G, Cambillau C: Structures of a legume lectin complexed with the human lactotransferrin N2 fragment, and with an isolated biantennary glycopeptide: role of the fucose moiety. Structure 1994, 2:209-219.
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Bourne Y, Rough P, Cambillau C: X-ray structure of a biantennary octasaccharide-lectin complex refined at 2.3/~, resolution. J Biol Chem 1992, 267:19?-203.
62.
Bourne Y, Bolgiano B, Liao DI, Strecker G, Cantau P, Herzberg O, Feizi T, Cambillau C: Crosslinking of mammalian lectin (galectin-1) by complex biantennary saccharides. Nat Struct Biol 1994, 1:863-870.
•
43. ••
Perez S, Mouhous-Riou N, Nifant'ev NE, Tsvetkov YE, Bachet B, Imberty A:Crystal and molecular structure of a histoblood group antigen involved in cell adhesion: the Lewis x trisaccharide. Glycobiology 1996, 6:537~542. The first description of the crystal structure of a histo-blood group carbohydrate determinant. Some interactions observed between adjacent trisaccharides can provide clues for carbohydrate-carbohydrate interactions in cell adhesion. 44.
YvelinF, Zhang YM, Mallet JM, Robert F, Jeannin Y, Sinay P: Crystal structure of the Lewis x trisaccharide. Carbohydr Lett 1996, 1:475-482.
45.
Kojima N, Fenderson BA, Stroud MR, Goldberg RI, Habermann R, Toyokuni T, Hakomori S: Further studies on cell adhesion based on LeX-Lex interaction, with new approaches: embryoglycan aggregation of F9 teratocarcinoma cells, and adhesion of various turnout cells based on Lex expression. G/ycoconj -I 1994, 11:238-248.
46.
Ni F: Complete relaxation analysis matrix of transferred nuclear Overhauser effects. J Magn Reson 1 992, 96:651-656.
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