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1.
Histochem Cell Biol ; 116(2): 183-93, 2001 Aug.
Article in English | MEDLINE | ID: mdl-11685546

ABSTRACT

The lectin from the mushroom Polysporus squamosus (PSL) has an extended carbohydrate combining site, which exhibits a high specificity and affinity toward the NeuAc5alpha2,6Galbeta1,4Glc/GlcNAc trisaccharide sequence of asparagine-linked oligosaccharides. Therefore, PSL should be a superior reagent to the lectin from Sambucus nigra (SNA), which does not discriminate between alpha2,6-linked NeuAc5 present either in asparagine- or serine/threonine-linked oligosaccharides. We have prepared a digoxigenin-conjugated PSL and applied it for histochemistry and blotting. We observed a more restricted staining pattern by PSL as compared to SNA in paraffin sections from different rat organs. Pretreatment of sections with N-glycanase F abolished PSL staining indicating that it interacts only with asparagine-linked oligosaccharides. Furthermore, PSL staining was neuraminidase sensitive. In contrast, SNA staining was only partially sensitive to N-glycanase F pretreatment demonstrating that it was in part due to alpha2,6-linked NeuAc5 present in serine/threonine-linked oligosaccharides. The most striking observation in this regard was that PSL, in contrast to SNA, did not stain the mucus of sheep submandibular gland, which is extremely rich in serine/threonine-linked Neu5Acalpha2,6N-acetylgalactosamine. Furthermore, in some tissues neuraminidase pretreatment resulted in increased intensity of SNA staining probably due to binding to exposed terminal N-acetylgalactosamine residues. Collectively, these results indicate that PSL is a useful tool for the histochemical detection of alpha2,6-linked NeuAc5 in asparagine-linked oligosaccharides.


Subject(s)
Agaricales/chemistry , Histocytochemistry/methods , Lectins/pharmacology , Oligosaccharides/analysis , Plant Lectins , Animals , Asparagine , Intestinal Mucosa/chemistry , Kidney/chemistry , Liver/chemistry , Rats , Rats, Sprague-Dawley , Ribosome Inactivating Proteins , Sheep , Submandibular Gland/chemistry
3.
Glycobiology ; 11(2): 141-7, 2001 Feb.
Article in English | MEDLINE | ID: mdl-11287401

ABSTRACT

The binding affinity and specificity of the mushroom Polyporus squamosus lectin has been determined by the recently developed method of frontal affinity chromatography coupled to electrospray mass spectrometry (FAC/MS). A micro-scale affinity column was prepared by immobilizing the lectin ( approximately 25 microg) onto porous glass beads in a tubing column (9.8 microl column volume). The column was then used to screen several oligosaccharide mixtures. The dissociation constants of 22 sialylated or sulfated oligosaccharides were evaluated against the immobilized lectin. The lectin was found to be highly specific for Neu5Acalpha2-6Galbeta1-4Glc/GlcNAc containing oligosaccharides with K(d) values near 10 microM. The FAC/MS assay permits the rapid determination of the dissociation constants of ligands as well as a higher throughput screening of compound mixtures, making it a valuable tool for affinity studies, especially for testing large numbers of compounds.


Subject(s)
Agaricales/metabolism , Chromatography, Affinity/methods , Lectins/metabolism , Spectrometry, Mass, Electrospray Ionization/methods , Binding Sites , Carbohydrate Sequence , Lectins/chemistry , Molecular Sequence Data , Protein Binding
4.
Eur J Biochem ; 268(9): 2609-15, 2001 May.
Article in English | MEDLINE | ID: mdl-11322880

ABSTRACT

Examination of lectins of banana (Musa acuminata) and the closely related plantain (Musa spp.) by the techniques of quantitative precipitation, hapten inhibition of precipitation, and isothermal titration calorimetry showed that they are mannose/glucose binding proteins with a preference for the alpha-anomeric form of these sugars. Both generate precipitin curves with branched chain alpha-mannans (yeast mannans) and alpha-glucans (glycogens, dextrans, and starches), but not with linear alpha-glucans containing only alpha1,4- and alpha1,6-glucosidic bonds (isolichenan and pullulan). The novel observation was made that banana and plantain lectins recognize internal alpha1,3-linked glucosyl residues, which occur in the linear polysaccharides elsinan and nigeran. Concanavalin A and lectins from pea and lentil, also mannose/glucose binding lectins, did not precipitate with any of these linear alpha-glucans. This is, the authors believe, the first report of the recognition of internal alpha1,3-glucosidic bonds by a plant lectin. It is possible that these lectins are present in the pulp of their respective fruit, complexed with starch.


Subject(s)
Carbohydrate Metabolism , Fruit/metabolism , Lectins/metabolism , Binding Sites , Carbohydrates/chemistry , Chemical Precipitation , Chromatography, Affinity , Fruit/chemistry , Lectins/chemistry , Lectins/isolation & purification , Plant Lectins
5.
Eur J Biochem ; 268(9): 2616-9, 2001 May.
Article in English | MEDLINE | ID: mdl-11322881

ABSTRACT

This paper extends our knowledge of the rather bizarre carbohydrate binding poperties of the banana lectin (Musa acuminata). Although a glucose/mannose binding protein which recognizes alpha-linked gluco-and manno-pyranosyl groups of polysaccharide chain ends, the banana lectin was shown to bind to internal 3-O-alpha-D-glucopyranosyl units. Now we report that this lectin also binds to the reducing glucosyl groups of beta-1,3-linked glucosyl oligosaccharides (e.g. laminaribiose oligomers). Additionally, banana lectin also recognizes beta1,6-linked glucosyl end groups (gentiobiosyl groups) as occur in many fungal beta1,3/1,6-linked polysaccharides. This behavior clearly distinguishes the banana lectin from other mannose/glucose binding lectins, such as concanavalin A and the pea, lentil and Calystegia sepium lectins.


Subject(s)
Carbohydrate Metabolism , Fruit/metabolism , Lectins/metabolism , Binding Sites , Carbohydrate Sequence , Carbohydrates/chemistry , Disaccharides/chemistry , Disaccharides/metabolism , Glucans/chemistry , Glucans/metabolism , Lectins/isolation & purification , Molecular Sequence Data , Oligosaccharides/chemistry , Oligosaccharides/metabolism , Plant Lectins
6.
Nucleic Acids Res ; 29(2): E4, 2001 Jan 15.
Article in English | MEDLINE | ID: mdl-11139634

ABSTRACT

Sephadex-binding RNA ligands (aptamers) were obtained through in vitro selection. They could be classified into two groups based on their consensus sequences and the aptamers from both groups showed strong binding to Sephadex G-100. One of the highest affinity aptamers, D8, was chosen for further characterization. Aptamer D8 bound to dextran B512, the soluble base material of Sephadex, but not to isomaltose, isomaltotriose and isomaltotetraose, suggesting that its optimal binding site might consist of more than four glucose residues linked via alpha-1,6 linkages. The aptamer was very specific to the Sephadex matrix and did not bind appreciably to other supporting matrices, such as Sepharose, Sephacryl, cellulose or pustulan. Using Sephadex G-100, the aptamer could be purified from a complex mixture of cellular RNA, giving an enrichment of at least 60 000-fold, compared with a non-specific control RNA. These RNA aptamers can be used as affinity tags for RNAs or RNA subunits of ribonucleoproteins to allow rapid purification from complex mixtures of RNA using only Sephadex.


Subject(s)
Dextrans/chemistry , RNA/chemistry , RNA/isolation & purification , Sepharose/analogs & derivatives , Acrylic Resins/chemistry , Binding, Competitive , Carbohydrate Sequence , Cellulose/chemistry , HeLa Cells , Humans , Ligands , Macromolecular Substances , Oligoribonucleotides/chemistry , Oligoribonucleotides/isolation & purification , Oligosaccharides/chemistry , Polysaccharides/chemistry , Sepharose/chemistry , Tumor Cells, Cultured
7.
J Biol Chem ; 275(14): 10623-9, 2000 Apr 07.
Article in English | MEDLINE | ID: mdl-10744758

ABSTRACT

A lectin has been purified from the carpophores of the mushroom Polyporus squamosus by a combination of affinity chromatography on beta-D-galactosyl-Synsorb and ion-exchange chromatography on DEAE-Sephacel. Gel filtration chromatography, SDS-polyacrylamide gel electrophoresis, and N-terminal amino acid sequencing indicated that the native lectin, designated P. squamosus agglutinin, is composed of two identical 28-kDa subunits associated by noncovalent bonds. P. squamosus agglutinin agglutinated human A, B, and O and rabbit red blood cells but precipitated only with human alpha(2)-macroglobulin, of many glycoproteins and polysaccharides tested. The detailed carbohydrate binding properties of the purified lectin were elucidated using three different approaches, i.e. precipitation inhibition assay (in solution binding assay), fluorescence quenching studies, and glycolipid binding by lectin staining on high-performance thin layer chromatography (solid-phase binding assay). Based on the results obtained by these assays, we conclude that although the P. squamosus lectin binds beta-D-galactosides, it has an extended carbohydrate-combining site that exhibits highest specificity and affinity toward nonreducing terminal Neu5Acalpha2, 6Galbeta1,4Glc/GlcNAc (6'-sialylated type II chain) of N-glycans (2000-fold stronger than toward galactose). The strict specificity of the lectin for alpha2,6-linked sialic acid renders this lectin a valuable tool for glycobiological studies in biomedical and cancer research.


Subject(s)
Agaricales/chemistry , Lectins/chemistry , Oligosaccharides/chemistry , ABO Blood-Group System , Amino Acids/analysis , Animals , Carbohydrate Conformation , Carbohydrate Sequence , Chromatography, Affinity , Chromatography, Gel , Chromatography, Ion Exchange , Glycosphingolipids/chemistry , Hemagglutination Tests , Humans , Lectins/isolation & purification , Molecular Sequence Data , Molecular Weight , Oligosaccharides/isolation & purification , Rabbits , Spores, Fungal/chemistry , alpha-Macroglobulins/chemistry
8.
J Biol Chem ; 274(47): 33300-5, 1999 Nov 19.
Article in English | MEDLINE | ID: mdl-10559206

ABSTRACT

An unusual lectin possessing two distinctly different types of carbohydrate-combining sites was purified from tubers of Xanthosoma sagittifolium L. by consecutive passage through two affinity columns, i.e. asialofetuin-Sepharose and invertase-Sepharose. SDS-polyacrylamide gel electrophoresis, N-terminal amino acid sequencing, and gel filtration chromatography of the purified lectin showed that the X. sagittifolium lectin is a heterotetrameric protein composed of four 12-kDa subunits (alpha(2)beta(2)) linked by noncovalent bonds. The results obtained by quantitative precipitation and hapten inhibition assays revealed that the lectin has two different types of carbohydrate-combining sites: one type for oligomannoses, which preferentially binds to a cluster of nonreducing terminal alpha1,3-linked mannosyl residues, and the other type for complex N-linked carbohydrates, which best accommodates a non-sialylated, triantennary oligosaccharide with N-acetyllactosamine (i.e. Galbeta1,4GlcNAc-) or lacto-N-biose (i.e. Galbeta1,3GlcNAc-) groups at its three nonreducing termini.


Subject(s)
Lectins/chemistry , Plants/chemistry , Carbohydrate Sequence , Lectins/isolation & purification , Molecular Sequence Data , Molecular Weight , Plant Lectins , Precipitin Tests , Protein Conformation
10.
Glycobiology ; 9(2): 173-9, 1999 Feb.
Article in English | MEDLINE | ID: mdl-9949194

ABSTRACT

The mannose/glucose-binding Dolichos lablab lectin (designated DLL) has been purified from seeds of Dolichos lablab (hyacinth bean) to electrophoretic homogeneity by affinity chromatography on an ovalbumin-Sepharose 4B column. The purified lectin gave a single symmetric protein peak with an apparent molecular mass of 67 kDa on gel filtration chromatography, and five bands ranging from 10 kDa to 22 kDa upon SDS-PAGE. N-Terminal sequence analysis of these bands revealed subunit heterogeneity due to posttranslational proteolytic truncation at different sites mostly at the carboxyl terminus. The carbohydrate binding properties of the purified lectin were investigated by three different approaches: hemagglutination inhibition assay, quantitative precipitation inhibition assay, and ELISA. On the basis of these studies, it is concluded that the Dolichos lablab lectin has neither an extended carbohydrate combining site, nor a hydrophobic binding site adjacent to it. The carbohydrate combining site of DLL appears to most effectively accommodate a nonreducing terminal alpha-d-mannosyl unit, and to be complementary to the C-3, C-4, and C-6 equatorial hydroxyl groups of alpha-d-mannopyranosyl and alpha-d-glucopyranosyl residues. DLL strongly precipitates murine IgM but not IgG, and the recent finding that this lectin interacts specifically with NIH 3T3 fibroblasts transfected with the Flt3 tyrosine kinase receptor and preserves human cord blood stem cells and progenitors in a quiescent state for prolonged periods in culture, make this lectin a valuable tool in biomedical research.


Subject(s)
Lectins/isolation & purification , Carbohydrate Metabolism , Chromatography, Affinity , Enzyme-Linked Immunosorbent Assay , Fabaceae/chemistry , Hemagglutination Inhibition Tests , Lectins/metabolism , Plant Lectins , Plants, Medicinal , Precipitin Tests , Protein Binding , Seeds/chemistry
11.
Biochemistry ; 37(48): 16952-7, 1998 Dec 01.
Article in English | MEDLINE | ID: mdl-9836588

ABSTRACT

The Griffonia simplicifolia-I (GS-I) isolectins have been used to probe the effect of lectin valence on their high-affinity binding to human erythrocytes. These tetrameric lectins are composed of A and B subunits and constitute a series of five isolectins (A4, A3B, A2B2, AB3, B4). The A subunit is specific for alpha-D-GalNAc end groups and binds to the blood type A determinant GalNAcalpha1, as well as to terminal alpha-D-Gal groups found on type B cells. The B subunit is specific for alpha-D-Gal end groups, and binds very specifically to type B erythrocytes. This series of isolectins is tetravalent (A4), trivalent (A3B), divalent (A2B2), and monovalent (AB3) for type A erythrocytes; thus, this system provides the opportunity to examine the effect of lectin valency on the association constants of these GS-I isolectins binding to cells. Cell binding experiments carried out using 125I-labeled GS-I isolectins and type A human erythrocytes allowed us to demonstrate that (1) the association constant of the isolectin monovalent for alpha-D-GalNAc (AB3) is virtually identical to its association constant for the haptenic sugar methyl-N-acetyl-alpha-D-galactosaminide, reported previously, and (2) the association constant of the GS-I isolectins for human type A erythrocytes increases with increasing valency of the isolectin. These results indicate that the increased affinity displayed by the GS-I isolectins for human type A erythrocytes is dependent on their multivalency, and not on an extended binding site nor on nonspecific, or noncarbohydrate, interactions of the lectin with the cell surface. These findings should be of general relevance to understanding the high-affinity interactions observed between other multivalent proteins and multivalent ligands (e.g., cell surfaces).


Subject(s)
ABO Blood-Group System , Erythrocytes/metabolism , Lectins/metabolism , Plant Lectins , Humans , Protein Binding
12.
Science ; 282(5391): 1049, 1998 Nov 06.
Article in English | MEDLINE | ID: mdl-9841450
13.
Eur J Biochem ; 254(2): 217-22, 1998 Jun 01.
Article in English | MEDLINE | ID: mdl-9660173

ABSTRACT

A cDNA library of Limax flavus was constructed and screened for sialic-acid-specific lectins. Complementary DNA clones were categorized into seven groups corresponding to closely related but different sequences. Group 1 clones contained an ORF encoding 199 amino acids including a sequence identical to the partial amino acid sequence obtained from the lectin protein. Within its 1074-bp 3' untranslated region, ten closely related 60-bp sequence repeats were found. Group 2 clones contained an ORF encoding a polypeptide chain of the same number of amino acid residues, with 89.1% overall identity to that of the group 1 and eight 60-bp repeat sequences in the 3' untranslated region. The remaining groups of clones contained ORF with highly similar full or partial sequences, with or without 60 bp repeats in the 3' untranslated region. The large number of closely related but different cDNA clones obtained indicated that the slug sialic-acid-specific lectin gene is a member of a multigene family. The lectin amino acid sequence showed significant similarity with the fibrinogen domain of human tenascin-C, with a human C-type serum lectin, and with pig ficolin. Immunostaining analysis of slug tissue for the lectin indicated that it is present primarily on the epidermal surface and in mucous glands. Recombinant slug lectin protein lacking the 20-amino-acid N-terminal signal sequence produced in a bacterial expression system from a group-1 clone accumulated as aggregates in inclusion bodies, suggesting that large-scale production of the active agglutinin may be possible.


Subject(s)
Lectins/genetics , Lectins/metabolism , Mollusca/genetics , Mollusca/metabolism , Amino Acid Sequence , Animals , Base Sequence , Cloning, Molecular , DNA, Complementary/genetics , Escherichia coli/genetics , Gene Expression , Hemagglutination , Humans , Inclusion Bodies/metabolism , Lectins/isolation & purification , Molecular Sequence Data , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , Sialic Acid Binding Immunoglobulin-like Lectins , Tissue Distribution
14.
Proc Natl Acad Sci U S A ; 95(10): 5462-7, 1998 May 12.
Article in English | MEDLINE | ID: mdl-9576904

ABSTRACT

Cell surface oligosaccharides have been shown to play essential biological roles in such diverse biological phenomena as cellular adhesion, molecular recognition, and inflammatory response. The development of high-affinity ligands capable of selectively recognizing a variety of small motifs in different oligosaccharides would be of significant interest as experimental and diagnostic tools. As a step toward this goal we have developed DNA ligands that recognize the disaccharide cellobiose, whether in soluble form or as the repeating unit of the polymer, cellulose. These DNA "aptamers" bind with high selectivity to cellobiose with little or no affinity for the related disaccharides lactose, maltose, and gentiobiose. Thus, the DNA ligands can discriminate sugar epimers, anomers, and disaccharide linkages.


Subject(s)
Cellobiose/metabolism , DNA/metabolism , Carbohydrate Conformation , Carbohydrate Sequence , Cellulose/metabolism , Gene Library , Kinetics , Ligands , Molecular Sequence Data , Sequence Analysis, DNA
15.
J Biol Chem ; 272(41): 25455-61, 1997 Oct 10.
Article in English | MEDLINE | ID: mdl-9325257

ABSTRACT

A unique mannose-binding lectin, highly specific for terminal Man(alpha1,3)Man groups, was isolated from bulbs of crocus (Crocus vernus All.). The lectin failed to bind to a mannose affinity column and was purified by simple gel permeation chromatography (Sephacryl S200). The purified lectin, obtained in crystalline form, had a molecular mass of 44 kDa on gel filtration and showed a single peptide band with a molecular mass of 11 kDa on SDS-polyacrylamide gel electrophoresis, indicating it to be a tetrameric protein composed of four identical subunits. The N-terminal amino acid sequence analysis of the crocus lectin showed essentially no homology with that of other mannose-binding bulb lectins. The crocus lectin selectively interacted with the wild type Saccharomyces cerevisiae and other mannans carrying terminal Man(alpha1,3)Man but not with those lacking this disaccharide unit. In hapten inhibition studies, methyl alpha-mannopyranoside did not inhibit the mannan-lectin interaction. Of various alpha-mannooligosaccharides, those having the Man(alpha1,3)Man sequence showed the highest inhibitory potency, confirming the strict requirement of lectin for terminal alpha1,3-linked mannosylmannose units. An affinity column of immobilized lectin enabled the complete resolution of yeast mannan and glycogen. The immobilized lectin may provide a useful tool for purification and analysis of biologically important polysaccharides and glycoproteins.


Subject(s)
Carrier Proteins/isolation & purification , Colchicum/chemistry , Disaccharides/metabolism , Lectins/isolation & purification , Plants, Medicinal , Carbohydrate Sequence , Carrier Proteins/chemistry , Chemical Precipitation , Glycoproteins/metabolism , Hydrogen-Ion Concentration , Lectins/chemistry , Mannans/metabolism , Molecular Sequence Data , Molecular Weight , Plant Lectins , Polysaccharides/metabolism , Saccharomyces cerevisiae
16.
Nat Struct Biol ; 4(10): 779-83, 1997 Oct.
Article in English | MEDLINE | ID: mdl-9334739

ABSTRACT

Amaranthus caudatus agglutinin contains a novel arrangement of four beta-trefoil domains. The sugar-binding site provides specificity for the carcinoma-associated T-antigen disaccharide even when 'masked' by other sugars.


Subject(s)
Antigens, Viral, Tumor/chemistry , Disaccharides , Lectins/chemistry , Protein Conformation , Benzyl Compounds , Crystallography, X-Ray/methods , Dimerization , Edible Grain , Models, Molecular , Plant Lectins , Ribosome Inactivating Proteins , Ribosome Inactivating Proteins, Type 1 , Seeds
17.
Eur J Biochem ; 245(3): 648-55, 1997 May 01.
Article in English | MEDLINE | ID: mdl-9183001

ABSTRACT

A second NeuAc(alpha2,6)Gal/GalNAc binding type 2 ribosome-inactivating protein (RIP), called SNAI' has been isolated from elderberry (Sambucus nigra) bark. SNAI' is a minor bark protein which closely resembles the previously described major Neu5Ac(alpha2,6)Gal/GalNAc binding type 2 RIP called SNAI with respect to its carbohydrate-binding specificity and ribosome-inactivating activity but has a different molecular structure. Molecular cloning revealed that the deduced amino acid sequence of SNAI' is highly similar to that of SNAI and that the difference in molecular structure between both proteins relies on a single cysteine residue present in the B chain of SNAI but absent from SNAI'. The isolation of SNAI' not only identifies a minor bark protein as a type 2 RIP but also further emphasizes the complexity of the type 2 RIP/lectin mixture present in the bark of elderberry.


Subject(s)
Lectins/isolation & purification , N-Glycosyl Hydrolases , Plant Proteins/isolation & purification , Plants/chemistry , Amino Acid Sequence , Cloning, Molecular , Lectins/genetics , Lectins/metabolism , Molecular Sequence Data , Molecular Structure , Plant Lectins , Plant Proteins/genetics , Plant Proteins/metabolism , Ribosome Inactivating Proteins , Ribosome Inactivating Proteins, Type 2 , Sequence Alignment
18.
Glycoconj J ; 14(2): 259-65, 1997 Feb.
Article in English | MEDLINE | ID: mdl-9111143

ABSTRACT

A novel plant lectin has been isolated from the rhizomes of Calystegia sepium (hedge bindweed) and partially characterized. The lectin is a dimeric protein composed of two identical non-covalently linked subunits of 16 kDa. Hapten inhibition studies indicate that the novel lectin is best inhibited by maltose and mannose and hence exhibits a sugar binding specificity that differs in some respects from that of all previously isolated plant lectins. Mitogenicity tests have shown that the Calystegia lectin is a powerful T-cell mitogen. Affinity purification of human, plant and fungal glycoproteins on immobilized C. sepium lectin demonstrates that this novel lectin can be used for the isolation of glycoconjugates from various sources. Moreover, it can be expected that by virtue of its distinct specificity, the new lectin will become an important tool in glycobiology.


Subject(s)
Carbohydrate Metabolism , Lectins/isolation & purification , Lectins/metabolism , Plants/chemistry , Agglutination , Agglutination Tests , Amino Acid Sequence , Amino Acids/analysis , Animals , Chromatography, Affinity/methods , Dose-Response Relationship, Drug , Erythrocytes/drug effects , Erythrocytes/metabolism , Fungal Proteins/chemistry , Fungal Proteins/metabolism , Glycoproteins/chemistry , Glycoproteins/metabolism , Hemagglutination , Humans , Lectins/pharmacology , Lymphocytes/drug effects , Mice , Mice, Inbred BALB C , Molecular Sequence Data , Plant Lectins , Plant Proteins/chemistry , Plant Proteins/metabolism , Rabbits , Spleen/drug effects , Substrate Specificity
19.
Glycoconj J ; 14(2): 281-8, 1997 Feb.
Article in English | MEDLINE | ID: mdl-9111146

ABSTRACT

A lectin was isolated from fruiting bodies of Agrocybe cylindracea by two ion-exchange chromatographies and gel filtration on Toyopearl HW55F. The lectin was homogeneous on polyacrylamide gel electrophoresis and its molecular mass was determined to be 30000 by gel filtration, and 15000 by sodium dodecylsulfate polyacrylamide gel electrophoresis, signifying a dimeric protein. Its carbohydrate-binding specificity was investigated both by sugar-hapten inhibition of hemagglutination and by enzyme-linked immunosorbent assay. The inhibition tests showed the affinity of the lectin to be weakly directed toward sialic acid and lactose, and the enhanced affinity toward trisaccharides containing the NeuAc alpha2,3Gal beta-structure. Importantly, the lectin strongly interacted with glycoconjugates containing NeuAc alpha2,3Gal beta1,3GlcNAc-/GalNAc sequences.


Subject(s)
Basidiomycota/chemistry , Carbohydrate Metabolism , Lectins/isolation & purification , Lectins/metabolism , Amino Acids/analysis , Animals , Carbohydrate Sequence , Carbohydrates/pharmacology , Caseins/metabolism , Enzyme-Linked Immunosorbent Assay/methods , Gangliosides/metabolism , Gangliosides/pharmacology , Glycoconjugates/chemistry , Glycoconjugates/metabolism , Glycoconjugates/pharmacology , Hemagglutination/drug effects , Laminin/metabolism , Lectins/pharmacology , Mice , Molecular Sequence Data , Molecular Weight , N-Acetylneuraminic Acid/metabolism , Oligosaccharides/pharmacology , Plant Proteins/isolation & purification , Plant Proteins/metabolism , Sequence Analysis , Serum Albumin, Bovine/chemistry , Serum Albumin, Bovine/metabolism , Substrate Specificity
20.
Glycoconj J ; 14(8): 889-96, 1997 Dec.
Article in English | MEDLINE | ID: mdl-9486421

ABSTRACT

A new mannose-binding lectin was isolated from Sternbergia lutea bulbs by affinity chromatography on an alpha(1-2)mannobiose-Synsorb column and purified further by gel filtration. This lectin (S. lutea agglutinin; SLA) appeared homogeneous by native-gel electrophoresis at pH 4.3, gel filtration chromatography on a Sephadex G-75 column, and SDS-polyacrylamide gel electrophoresis, These data indicate that SLA is a dimeric protein (20 kDa) composed of two identical subunits of 10 kDa which are linked by non-covalent interactions. The carbohydrate binding specificity of the lectin was investigated by quantitative precipitation and hapten inhibition assays. It is an alpha-D-mannose-specific lectin that interacts to form precipitates with various alpha-mannans, galactomannan and asialo-thyroglobulin, but not with alpha-glucans and thyroglobulin. Of the monosaccharides tested only D-mannose was a hapten inhibitor of the SLA-asialothyroglobulin precipitation system, whereas D-glucose, D-galactose and L-arabinose were not. The lectin appears to be highly specific for terminal alpha(1-3)-mannooligosaccharides. The primary structure of SLA appears to be quite similar to that of the snow drop (Galanthus nivalis) bulb lectin which is a mannose-binding lectin from the same plant family Amaryllidaceae. The N-terminal 46 amino acid sequence SLA showed 76% homology with that of GNA.


Subject(s)
Carrier Proteins/isolation & purification , Carrier Proteins/metabolism , Lectins/isolation & purification , Lectins/metabolism , Mannose-Binding Lectins , Amino Acid Sequence , Binding Sites , Carbohydrate Conformation , Carbohydrate Sequence , Carrier Proteins/genetics , Collectins , Dimerization , Galanthus , Lectins/genetics , Molecular Sequence Data , Molecular Weight , Oligosaccharides/chemistry , Oligosaccharides/metabolism , Plant Lectins , Plants/chemistry , Plants/genetics , Protein Conformation , Sequence Homology, Amino Acid
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