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Remy Loris - One of the best experts on this subject based on the ideXlab platform.

  • interplay between metal binding and cis trans isomerization in Legume Lectins structural and thermodynamic study of p angolensis Lectin
    Journal of Molecular Biology, 2006
    Co-Authors: Abel Garciapino, Lode Wyns, L Buts, Remy Loris
    Abstract:

    The interplay between metal binding, carbohydrate binding activity, stability and structure of the Lectin from Pterocarpus angolensis was investigated. Removal of the metals leads to a more flexible form of the protein with significantly less conformational stability. Crystal structures of this metal-free form show significant structural rearrangements, although some structural features that allow the binding of sugars are retained. We propose that substitution of an asparagine residue at the start of the C-terminal β-strand of the Legume Lectin monomer hinders the trans-isomerization of the cis-peptide bond upon demetallization and constitutes an intramolecular switch governing the isomer state of the non-proline bond and ultimately the Lectin phenotype.

  • Interplay between metal binding and cis/trans isomerization in Legume Lectins: structural and thermodynamic study of P. angolensis Lectin.
    Journal of Molecular Biology, 2006
    Co-Authors: Abel Garcia-pino, Lode Wyns, L Buts, Remy Loris
    Abstract:

    The interplay between metal binding, carbohydrate binding activity, stability and structure of the Lectin from Pterocarpus angolensis was investigated. Removal of the metals leads to a more flexible form of the protein with significantly less conformational stability. Crystal structures of this metal-free form show significant structural rearrangements, although some structural features that allow the binding of sugars are retained. We propose that substitution of an asparagine residue at the start of the C-terminal β-strand of the Legume Lectin monomer hinders the trans-isomerization of the cis-peptide bond upon demetallization and constitutes an intramolecular switch governing the isomer state of the non-proline bond and ultimately the Lectin phenotype.

  • weak protein protein interactions in Lectins the crystal structure of a vegetative Lectin from the Legume dolichos biflorus
    Journal of Molecular Biology, 2001
    Co-Authors: L Buts, Lode Wyns, Marilynn E. Etzler, Remy Loris, Minhhoa Daothi, Thomas Hamelryck
    Abstract:

    Abstract The Legume Lectins are widely used as a model system for studying protein-carbohydrate and protein-protein interactions. They exhibit a fascinating quaternary structure variation, which becomes important when they interact with multivalent glycoconjugates, for instance those on cell surfaces. Recently, it has become clear that certain Lectins form weakly associated oligomers. This phenomenon may play a role in the regulation of receptor crosslinking and subsequent signal transduction. The crystal structure of DB58, a dimeric Lectin from the Legume Dolichos biflorus reveals a separate dimer of a previously unobserved type, in addition to a tetramer consisting of two such dimers. This tetramer resembles that formed by DBL, the seed Lectin from the same plant. A single amino acid substitution in DB58 affects the conformation and flexibility of a loop in the canonical dimer interface. This disrupts the formation of a stable DBL-like tetramer in solution, but does not prohibit its formation in suitable conditions, which greatly increases the possibilities for the crosslinking of multivalent ligands. The non-canonical DB58 dimer has a buried symmetrical α helix, which can be present in the crystal in either of two antiparallel orientations. Two existing structures and datasets for Lectins with similar quaternary structures were reconsidered. A central α helix could be observed in the soybean Lectin, but not in the leucoagglutinating Lectin from Phaseolus vulgaris. The relative position and orientation of the carbohydrate-binding sites in the DB58 dimer may affect its ability to crosslink mulitivalent ligands, compared to the other Legume Lectin dimers.

  • Structural basis of carbohydrate recognition by Lectin II from Ulex europaeus, a protein with a promiscuous carbohydrate-binding site.
    Journal of Molecular Biology, 2000
    Co-Authors: Remy Loris, Anne Imberty, H. De Greve, Joris Messens, Lode Wyns
    Abstract:

    Protein-carbohydrate interactions are the language of choice for inter-cellular communication. The Legume Lectins form a large family of homologous proteins that exhibit a wide variety of carbohydrate specificities. The Legume Lectin family is therefore highly suitable as a model system to study the structural principles of protein-carbohydrate recognition. Until now, structural data are only available for two specificity families: Man/Glc and Gal/GalNAc. No structural data are available for any of the fucose or chitobiose specific Lectins. The crystal structure of Ulex europaeus (UEA-II) is the first of a Legume Lectin belonging to the chitobiose specificity group. The complexes with N-acetylglucosamine, galactose and fucosylgalactose show a promiscuous primary binding site capable of accommodating both N-acetylglucos amine or galactose in the primary binding site. The hydrogen bonding network in these complexes can be considered suboptimal, in agreement with the low affinities of these sugars. In the complexes with chitobiose, lactose and fucosyllactose this suboptimal hydrogen bonding network is compensated by extensive hydrophobic interactions in a Glc/GlcNAc binding subsite. UEA-II thus forms the first example of a Legume Lectin with a promiscuous binding site and illustrates the importance of hydrophobic interactions in protein-carbohydrate complexes. Together with other known Legume Lectin crystal structures, it shows how different specificities can be grafted upon a conserved structural framework.

  • Carbohydrate binding, quaternary structure and a novel hydrophobic binding site in two Legume Lectin oligomers from Dolichos biflorus.
    Journal of Molecular Biology, 1999
    Co-Authors: Thomas Hamelryck, Lode Wyns, Elias Fernandez, Gérard Strecker, Julie Bouckaert, Remy Loris, Anne Imberty, Marilynn E. Etzler
    Abstract:

    Abstract The seed Lectin (DBL) from the leguminous plant Dolichos biflorus has a unique specificity among the members of the Legume Lectin family because of its high preference for GalNAc over Gal. In addition, precipitation of blood group A+H substance by DBL is slightly better inhibited by a blood group A trisaccharide (GalNAc(α1-3)[Fuc(α1-2)]Gal) containing pentasaccharide, and about 40 times better by the Forssman disaccharide (GalNAc(α1-3)GalNAc) than by GalNAc. We report the crystal structures of the DBL-blood group A trisaccharide complex and the DBL-Forssman disaccharide complex. A comparison with the binding sites of Gal-binding Legume Lectins indicates that the low affinity of DBL for Gal is due to the substitution of a conserved aromatic residue by an aliphatic residue (Leu127). Binding studies with a Leu127Phe mutant corroborate these conclusions. DBL has a higher affinity for GalNAc because the N-acetyl group compensates for the loss of aromatic stacking in DBL by making a hydrogen bond with the backbone amide group of Gly103 and a hydrophobic contact with the side-chains of Trp132 and Tyr104. Some Legume Lectins possess a hydrophobic binding site that binds adenine and adenine-derived plant hormones, i.e. cytokinins. The exact function of this binding site is unknown, but adenine/cytokinin-binding Legume Lectins might be involved in storage of plant hormones or plant growth regulation. The structures of DBL in complex with adenine and of the dimeric stem and leaf Lectin (DB58) from the same plant provide the first structural data on these binding sites. Both oligomers possess an unusual architecture, featuring an α-helix sandwiched between two monomers. In both oligomers, this α-helix is directly involved in the formation of the hydrophobic binding site. DB58 adopts a novel quaternary structure, related to the quaternary structure of the DBL heterotetramer, and brings the number of know Legume Lectin dimer types to four.

Avadhesha Surolia - One of the best experts on this subject based on the ideXlab platform.

  • Impact of glycosylation on stability, structure and unfolding of soybean agglutinin (SBA): an insight from thermal perturbation molecular dynamics simulations
    Glycoconjugate Journal, 2015
    Co-Authors: Swagata Halder, Avadhesha Surolia, Chaitali Mukhopadhyay
    Abstract:

    Glycosylation has been recognized as one of the most prevalent and complex post-translational modification s of proteins involving numerous enzymes and substrates. Its effect on the protein conformational transitions is not clearly understood yet. In this study, we have examined the effect of glycosylation on protein stability using molecular dynamics simulation of Legume Lectin soybean agglutinin (SBA). Its glycosylated moiety consists of high mannose type N-linked glycan (Man_9GlcNAc_2). To unveil the structural perturbations during thermal unfolding of these two forms, we have studied and compared them to the experimental results. From the perspective of dynamics, our simulations revealed that the nonglycosylated monomeric form is less stable than corresponding glycosylated form at normal and elevated temperatures. Moreover, at elevated temperature thermal destabilization is more prominent in solvent exposed loops, turns and ends of distinct β sheets. SBA maintains it folded structure due to some important saltbridges, hydrogen bonds and hydrophobic interactions within the protein. The reducing terminal GlcNAc residues interact with the protein residues VAL161, PRO182 and SER225 via hydrophobic and via hydrogen bonding with ASN 9 and ASN 75. Our simulations also revealed that single glycosylation (ASN75) has no significant effect on corresponding cis peptide angle orientation. This atomistic description might have important implications for understanding the functionality and stability of Soybean agglutinin.

  • REVIEW ARTICLE Insights into the quaternary association of proteins through structure graphs: a case study of Lectins
    2015
    Co-Authors: K. V. Brinda, Avadhesha Surolia, Sarawathi Vishveshwara
    Abstract:

    The unique three-dimensional structure of both monomeric and oligomeric proteins is encoded in their sequence. The biological functions of proteins are dependent on their tertiary and quater-nary structures, and hence it is important to understand the deter-minants of quaternary association in proteins. Although a large number of investigations have been carried out in this direction, the underlying principles of protein oligomerization are yet to be completely understood. Recently, new insights into this problem have been gained from the analysis of structure graphs of proteins belonging to the Legume Lectin family. The Legume Lectins are an interesting family of proteins with very similar tertiary structures but varied quaternary structures. Hence they have become a very good model with which to analyse the role of primary struc-tures in determining the modes of quaternary association. The present review summarizes the results of a Legume Lectin study a

  • Insights into the quaternary association of proteins through structure graphs: a case study of Lectins.
    Biochemical Journal, 2005
    Co-Authors: K. V. Brinda, Avadhesha Surolia, Sarawathi Vishveshwara
    Abstract:

    The unique three-dimensional structure of both monomeric and oligomeric proteins is encoded in their sequence. The biological functions of proteins are dependent on their tertiary and quaternary structures, and hence it is important to understand the determinants of quaternary association in proteins. Although a large number of investigations have been carried out in this direction, the underlying principles of protein oligomerization are yet to be completely understood. Recently, new insights into this problem have been gained from the analysis of structure graphs of proteins belonging to the Legume Lectin family. The Legume Lectins are an interesting family of proteins with very similar tertiary structures but varied quaternary structures. Hence they have become a very good model with which to analyse the role of primary structures in determining the modes of quaternary association. The present review summarizes the results of a Legume Lectin study as well as those obtained from a similar analysis carried out here on the animal Lectins, namely gaLectins, pentraxins, calnexin, calreticulin and rhesus rotavirus Vp4 sialic-acid-binding domain. The Lectin structure graphs have been used to obtain clusters of non-covalently interacting amino acid residues at the intersubunit interfaces. The present study, performed along with traditional sequence alignment methods, has provided the signature sequence motifs for different kinds of quaternary association seen in Lectins. Furthermore, the network representation of the Lectin oligomers has enabled us to detect the residues which make extensive interactions (‘hubs’) across the oligomeric interfaces that can be targetted for interface-destabilizing mutations. The present review also provides an overview of the methodology involved in representing oligomeric protein structures as connected networks of amino acid residues. Further, it illustrates the potential of such a representation in elucidating the structural determinants of protein–protein association in general and will be of significance to protein chemists and structural biologists.

  • conformational stability of Legume Lectins reflect their different modes of quaternary association solvent denaturation studies on concanavalin a and winged bean acidic agglutinin
    Biochemistry, 2002
    Co-Authors: Nivedita Mitra, V R Srinivas, Nisar Ahmad, T N C Ramya, Bhanuprakash G Reddy, Avadhesha Surolia
    Abstract:

    Thermodynamic parameters associated with the unfolding of the Legume Lectin, WBA II, were determined by isothermal denaturation. The analysis of isothermal denaturation data provided values for conformational stability and heat capacity for WBA II unfolding. To explore the role of intersubunit contact in stability, we carried out similar studies under identical conditions on Concanavalin A, a Legume Lectin of nearly similar size, buried hydrophobic surface area and tertiary structure to that of WBA II but with a different oligomerization pattern. Both proteins showed a reversible two-state unfolding with guanidine hydrochloride. As expected, the change in heat capacity upon unfolding was similar for both proteins at 3.5 and 3.7 kcal $mol^-^1 K^-^1$ for Concanavalin A and WBA II, respectively. Although the $\bigtriangleup G_{H20}$ at the maximum stability of both proteins is around 16 kcal/mol, Concanavalin A exhibits greater stability at higher temperatures. The Tg obtained for Concanavalin A and WBA II were $21 ^\circ C$ apart at 87.2 and $66.6 ^\circ C$, respectively. The higher conformational stability at higher temperatures and the Tg of Concanavalin A as compared to that of WBA II are largely due to substantial differences in the degree of subunit contact in these dimeric proteins. Ionic interactions and hydrogen bonding between the monomers of the two proteins also seem to play a significant role in the observed stability differences between these two proteins.

  • Legume Lectins a paradigm in quaternary structure variations arising from similar tertiary structural fold
    2002
    Co-Authors: Nivedita Mitra, V R Srinivas, Avadhesha Surolia
    Abstract:

    Proteins belonging to the Legume Lectin family are characterized by similarity in their tertiary structures and differences in their modes of quatemary association which are brought about by minor variations in amino acid sequences. It is observed that differences in the modes of their association cause them to adopt different stabilities and folding patterns as studied by differential scanning calorimetry and isothemwal denaturation experiments. The proteins with the canonical and the handshake mode of association show a two-state denaturation profile but differ in their calorimetric to van't Hoff enthalpy ratio, thus indicating that these proteins unfold in slightly different ways. In this class of proteins we also see a homotetrameric protein like the peanut agglutinin, which neither shows a D 2 nor a four-fold symmetry This kind of unusual structure confers upon it an unusual folding pathway where the presence of a partially unfolded and active monomeric intermediate is observed. Thus we see that these proteins are good specimens to study the effects of minor alterations in their sequences on their oligomeric association.

Lode Wyns - One of the best experts on this subject based on the ideXlab platform.

  • interplay between metal binding and cis trans isomerization in Legume Lectins structural and thermodynamic study of p angolensis Lectin
    Journal of Molecular Biology, 2006
    Co-Authors: Abel Garciapino, Lode Wyns, L Buts, Remy Loris
    Abstract:

    The interplay between metal binding, carbohydrate binding activity, stability and structure of the Lectin from Pterocarpus angolensis was investigated. Removal of the metals leads to a more flexible form of the protein with significantly less conformational stability. Crystal structures of this metal-free form show significant structural rearrangements, although some structural features that allow the binding of sugars are retained. We propose that substitution of an asparagine residue at the start of the C-terminal β-strand of the Legume Lectin monomer hinders the trans-isomerization of the cis-peptide bond upon demetallization and constitutes an intramolecular switch governing the isomer state of the non-proline bond and ultimately the Lectin phenotype.

  • Interplay between metal binding and cis/trans isomerization in Legume Lectins: structural and thermodynamic study of P. angolensis Lectin.
    Journal of Molecular Biology, 2006
    Co-Authors: Abel Garcia-pino, Lode Wyns, L Buts, Remy Loris
    Abstract:

    The interplay between metal binding, carbohydrate binding activity, stability and structure of the Lectin from Pterocarpus angolensis was investigated. Removal of the metals leads to a more flexible form of the protein with significantly less conformational stability. Crystal structures of this metal-free form show significant structural rearrangements, although some structural features that allow the binding of sugars are retained. We propose that substitution of an asparagine residue at the start of the C-terminal β-strand of the Legume Lectin monomer hinders the trans-isomerization of the cis-peptide bond upon demetallization and constitutes an intramolecular switch governing the isomer state of the non-proline bond and ultimately the Lectin phenotype.

  • weak protein protein interactions in Lectins the crystal structure of a vegetative Lectin from the Legume dolichos biflorus
    Journal of Molecular Biology, 2001
    Co-Authors: L Buts, Lode Wyns, Marilynn E. Etzler, Remy Loris, Minhhoa Daothi, Thomas Hamelryck
    Abstract:

    Abstract The Legume Lectins are widely used as a model system for studying protein-carbohydrate and protein-protein interactions. They exhibit a fascinating quaternary structure variation, which becomes important when they interact with multivalent glycoconjugates, for instance those on cell surfaces. Recently, it has become clear that certain Lectins form weakly associated oligomers. This phenomenon may play a role in the regulation of receptor crosslinking and subsequent signal transduction. The crystal structure of DB58, a dimeric Lectin from the Legume Dolichos biflorus reveals a separate dimer of a previously unobserved type, in addition to a tetramer consisting of two such dimers. This tetramer resembles that formed by DBL, the seed Lectin from the same plant. A single amino acid substitution in DB58 affects the conformation and flexibility of a loop in the canonical dimer interface. This disrupts the formation of a stable DBL-like tetramer in solution, but does not prohibit its formation in suitable conditions, which greatly increases the possibilities for the crosslinking of multivalent ligands. The non-canonical DB58 dimer has a buried symmetrical α helix, which can be present in the crystal in either of two antiparallel orientations. Two existing structures and datasets for Lectins with similar quaternary structures were reconsidered. A central α helix could be observed in the soybean Lectin, but not in the leucoagglutinating Lectin from Phaseolus vulgaris. The relative position and orientation of the carbohydrate-binding sites in the DB58 dimer may affect its ability to crosslink mulitivalent ligands, compared to the other Legume Lectin dimers.

  • Structural basis of carbohydrate recognition by Lectin II from Ulex europaeus, a protein with a promiscuous carbohydrate-binding site.
    Journal of Molecular Biology, 2000
    Co-Authors: Remy Loris, Anne Imberty, H. De Greve, Joris Messens, Lode Wyns
    Abstract:

    Protein-carbohydrate interactions are the language of choice for inter-cellular communication. The Legume Lectins form a large family of homologous proteins that exhibit a wide variety of carbohydrate specificities. The Legume Lectin family is therefore highly suitable as a model system to study the structural principles of protein-carbohydrate recognition. Until now, structural data are only available for two specificity families: Man/Glc and Gal/GalNAc. No structural data are available for any of the fucose or chitobiose specific Lectins. The crystal structure of Ulex europaeus (UEA-II) is the first of a Legume Lectin belonging to the chitobiose specificity group. The complexes with N-acetylglucosamine, galactose and fucosylgalactose show a promiscuous primary binding site capable of accommodating both N-acetylglucos amine or galactose in the primary binding site. The hydrogen bonding network in these complexes can be considered suboptimal, in agreement with the low affinities of these sugars. In the complexes with chitobiose, lactose and fucosyllactose this suboptimal hydrogen bonding network is compensated by extensive hydrophobic interactions in a Glc/GlcNAc binding subsite. UEA-II thus forms the first example of a Legume Lectin with a promiscuous binding site and illustrates the importance of hydrophobic interactions in protein-carbohydrate complexes. Together with other known Legume Lectin crystal structures, it shows how different specificities can be grafted upon a conserved structural framework.

  • Carbohydrate binding, quaternary structure and a novel hydrophobic binding site in two Legume Lectin oligomers from Dolichos biflorus.
    Journal of Molecular Biology, 1999
    Co-Authors: Thomas Hamelryck, Lode Wyns, Elias Fernandez, Gérard Strecker, Julie Bouckaert, Remy Loris, Anne Imberty, Marilynn E. Etzler
    Abstract:

    Abstract The seed Lectin (DBL) from the leguminous plant Dolichos biflorus has a unique specificity among the members of the Legume Lectin family because of its high preference for GalNAc over Gal. In addition, precipitation of blood group A+H substance by DBL is slightly better inhibited by a blood group A trisaccharide (GalNAc(α1-3)[Fuc(α1-2)]Gal) containing pentasaccharide, and about 40 times better by the Forssman disaccharide (GalNAc(α1-3)GalNAc) than by GalNAc. We report the crystal structures of the DBL-blood group A trisaccharide complex and the DBL-Forssman disaccharide complex. A comparison with the binding sites of Gal-binding Legume Lectins indicates that the low affinity of DBL for Gal is due to the substitution of a conserved aromatic residue by an aliphatic residue (Leu127). Binding studies with a Leu127Phe mutant corroborate these conclusions. DBL has a higher affinity for GalNAc because the N-acetyl group compensates for the loss of aromatic stacking in DBL by making a hydrogen bond with the backbone amide group of Gly103 and a hydrophobic contact with the side-chains of Trp132 and Tyr104. Some Legume Lectins possess a hydrophobic binding site that binds adenine and adenine-derived plant hormones, i.e. cytokinins. The exact function of this binding site is unknown, but adenine/cytokinin-binding Legume Lectins might be involved in storage of plant hormones or plant growth regulation. The structures of DBL in complex with adenine and of the dimeric stem and leaf Lectin (DB58) from the same plant provide the first structural data on these binding sites. Both oligomers possess an unusual architecture, featuring an α-helix sandwiched between two monomers. In both oligomers, this α-helix is directly involved in the formation of the hydrophobic binding site. DB58 adopts a novel quaternary structure, related to the quaternary structure of the DBL heterotetramer, and brings the number of know Legume Lectin dimer types to four.

Chaitali Mukhopadhyay - One of the best experts on this subject based on the ideXlab platform.

  • Impact of glycosylation on stability, structure and unfolding of soybean agglutinin (SBA): an insight from thermal perturbation molecular dynamics simulations
    Glycoconjugate Journal, 2015
    Co-Authors: Swagata Halder, Avadhesha Surolia, Chaitali Mukhopadhyay
    Abstract:

    Glycosylation has been recognized as one of the most prevalent and complex post-translational modification s of proteins involving numerous enzymes and substrates. Its effect on the protein conformational transitions is not clearly understood yet. In this study, we have examined the effect of glycosylation on protein stability using molecular dynamics simulation of Legume Lectin soybean agglutinin (SBA). Its glycosylated moiety consists of high mannose type N-linked glycan (Man_9GlcNAc_2). To unveil the structural perturbations during thermal unfolding of these two forms, we have studied and compared them to the experimental results. From the perspective of dynamics, our simulations revealed that the nonglycosylated monomeric form is less stable than corresponding glycosylated form at normal and elevated temperatures. Moreover, at elevated temperature thermal destabilization is more prominent in solvent exposed loops, turns and ends of distinct β sheets. SBA maintains it folded structure due to some important saltbridges, hydrogen bonds and hydrophobic interactions within the protein. The reducing terminal GlcNAc residues interact with the protein residues VAL161, PRO182 and SER225 via hydrophobic and via hydrogen bonding with ASN 9 and ASN 75. Our simulations also revealed that single glycosylation (ASN75) has no significant effect on corresponding cis peptide angle orientation. This atomistic description might have important implications for understanding the functionality and stability of Soybean agglutinin.

Thomas Hamelryck - One of the best experts on this subject based on the ideXlab platform.

  • weak protein protein interactions in Lectins the crystal structure of a vegetative Lectin from the Legume dolichos biflorus
    Journal of Molecular Biology, 2001
    Co-Authors: L Buts, Lode Wyns, Marilynn E. Etzler, Remy Loris, Minhhoa Daothi, Thomas Hamelryck
    Abstract:

    Abstract The Legume Lectins are widely used as a model system for studying protein-carbohydrate and protein-protein interactions. They exhibit a fascinating quaternary structure variation, which becomes important when they interact with multivalent glycoconjugates, for instance those on cell surfaces. Recently, it has become clear that certain Lectins form weakly associated oligomers. This phenomenon may play a role in the regulation of receptor crosslinking and subsequent signal transduction. The crystal structure of DB58, a dimeric Lectin from the Legume Dolichos biflorus reveals a separate dimer of a previously unobserved type, in addition to a tetramer consisting of two such dimers. This tetramer resembles that formed by DBL, the seed Lectin from the same plant. A single amino acid substitution in DB58 affects the conformation and flexibility of a loop in the canonical dimer interface. This disrupts the formation of a stable DBL-like tetramer in solution, but does not prohibit its formation in suitable conditions, which greatly increases the possibilities for the crosslinking of multivalent ligands. The non-canonical DB58 dimer has a buried symmetrical α helix, which can be present in the crystal in either of two antiparallel orientations. Two existing structures and datasets for Lectins with similar quaternary structures were reconsidered. A central α helix could be observed in the soybean Lectin, but not in the leucoagglutinating Lectin from Phaseolus vulgaris. The relative position and orientation of the carbohydrate-binding sites in the DB58 dimer may affect its ability to crosslink mulitivalent ligands, compared to the other Legume Lectin dimers.

  • Carbohydrate binding, quaternary structure and a novel hydrophobic binding site in two Legume Lectin oligomers from Dolichos biflorus.
    Journal of Molecular Biology, 1999
    Co-Authors: Thomas Hamelryck, Lode Wyns, Elias Fernandez, Gérard Strecker, Julie Bouckaert, Remy Loris, Anne Imberty, Marilynn E. Etzler
    Abstract:

    Abstract The seed Lectin (DBL) from the leguminous plant Dolichos biflorus has a unique specificity among the members of the Legume Lectin family because of its high preference for GalNAc over Gal. In addition, precipitation of blood group A+H substance by DBL is slightly better inhibited by a blood group A trisaccharide (GalNAc(α1-3)[Fuc(α1-2)]Gal) containing pentasaccharide, and about 40 times better by the Forssman disaccharide (GalNAc(α1-3)GalNAc) than by GalNAc. We report the crystal structures of the DBL-blood group A trisaccharide complex and the DBL-Forssman disaccharide complex. A comparison with the binding sites of Gal-binding Legume Lectins indicates that the low affinity of DBL for Gal is due to the substitution of a conserved aromatic residue by an aliphatic residue (Leu127). Binding studies with a Leu127Phe mutant corroborate these conclusions. DBL has a higher affinity for GalNAc because the N-acetyl group compensates for the loss of aromatic stacking in DBL by making a hydrogen bond with the backbone amide group of Gly103 and a hydrophobic contact with the side-chains of Trp132 and Tyr104. Some Legume Lectins possess a hydrophobic binding site that binds adenine and adenine-derived plant hormones, i.e. cytokinins. The exact function of this binding site is unknown, but adenine/cytokinin-binding Legume Lectins might be involved in storage of plant hormones or plant growth regulation. The structures of DBL in complex with adenine and of the dimeric stem and leaf Lectin (DB58) from the same plant provide the first structural data on these binding sites. Both oligomers possess an unusual architecture, featuring an α-helix sandwiched between two monomers. In both oligomers, this α-helix is directly involved in the formation of the hydrophobic binding site. DB58 adopts a novel quaternary structure, related to the quaternary structure of the DBL heterotetramer, and brings the number of know Legume Lectin dimer types to four.

  • Legume Lectin structure.
    Biochimica et Biophysica Acta, 1998
    Co-Authors: Remy Loris, Thomas Hamelryck, Julie Bouckaert, Lode Wyns
    Abstract:

    Abstract The Legume Lectins are a large family of homologous carbohydrate binding proteins that are found mainly in the seeds of most Legume plants. Despite their strong similarity on the level of their amino acid sequences and tertiary structures, their carbohydrate specificities and quaternary structures vary widely. In this review we will focus on the structural features of Legume Lectins and their complexes with carbohydrates. These will be discussed in the light of recent mutagenesis results when appropriate. Monosaccharide specificity seems to be achieved by the use of a conserved core of residues that hydrogen bond to the sugar, and a variable loop that determines the exact shape of the monosaccharide binding site. The higher affinity for particular oligosaccharides and monosaccharides containing a hydrophobic aglycon results mainly from a few distinct subsites next to the monosaccharide binding site. These subsites consist of a small number of variable residues and are found in both the mannose and galactose specificity groups. The quaternary structures of these proteins form the basis of a higher level of specificity, where the spacing between individual epitopes of multivalent carbohydrates becomes important. This results in homogeneous cross-linked lattices even in mixed precipitation systems, and is of relevance for their effects on the biological activities of cells such as mitogenic responses. Quaternary structure is also thought to play an important role in the high affinity interaction between some Legume Lectins and adenine and a series of adenine-derived plant hormones. The molecular basis of the variation in quaternary structure in this group of proteins is poorly understood.

  • Crystal structure of arcelin-5, a Lectin-like defense protein from Phaseolus vulgaris
    Journal of Biological Chemistry, 1996
    Co-Authors: Thomas Hamelryck, Lode Wyns, F. Poortmans, Geert Angenon, Alain Goossens, Marc Van Montagu, Remy Loris
    Abstract:

    Abstract In the seeds of the Legume plants, a class of sugar-binding proteins with high structural and sequential identity is found, generally called the Legume Lectins. The seeds of the common bean (Phaseolus vulgaris) contain, besides two such Lectins, a Lectin-like defense protein called arcelin, in which one sugar binding loop is absent. Here we report the crystal structure of arcelin-5 (Arc5), one of the electrophoretic variants of arcelin, solved at a resolution of 2.7 A. The R factor of the refined structure is 20.6%, and the free R factor is 27.1%. The main difference between Arc5 and the Legume Lectins is the absence of the metal binding loop. The bound metals are necessary for the sugar binding capabilities of the Legume Lectins and stabilize an Ala-Asp cis-peptide bond. Surprisingly, despite the absence of the metal binding site in Arc5, this cis-peptide bond found in all Legume Lectin structures is still present, although the Asp residue has been replaced by a Tyr residue. Despite the high identity between the different Legume Lectin sequences, they show a broad range of quaternary structures. The structures of three different dimers and three different tetramers have been solved. Arc5 crystallized as a monomer, bringing the number of known quaternary structures to seven.

  • The Crystallographic Structure of Phytohemagglutinin-L
    Journal of Biological Chemistry, 1996
    Co-Authors: Thomas Hamelryck, Lode Wyns, F. Poortmans, Maarten J Chrispeels, Remy Loris
    Abstract:

    Abstract The structure of phytohemagglutinin-L (PHA-L), a leucoagglutinating seed Lectin from Phaseolus vulgaris, has been solved with molecular replacement using the coordinates of lentil Lectin as model, and refined at a resolution of 2.8 A. The final R-factor of the structure is 20.0%. The quaternary structure of the PHA-L tetramer differs from the structures of the concanavalin A and peanut Lectin tetramers, but resembles the structure of the soybean agglutinin tetramer. PHA-L consists of two canonical Legume Lectin dimers that pack together through the formation of a close contact between two β-strands. Of the two covalently bound oligosaccharides per monomer, only one GlcNAc residue per monomer is visible in the electron density. In this article we describe the structure of PHA-L, and we discuss the putative position of the high affinity adenine-binding site present in a number of Legume Lectins. A comparison with transthyretin, a protein that shows a remarkable resemblance to PHA-L, gives further ground to our proposal.