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1.
Imberty A Gautier C Lescar J Pérez S Wyns L Loris R 《The Journal of biological chemistry》2000,275(23):17541-17548
Seeds from the legume tree Maackia amurensis contain two lectins that can agglutinate different blood cell types. Their specificity toward sialylated oligosaccharides is unique among legume lectins; the leukoagglutinin preferentially binds to sialyllactosamine (alphaNeuAc(2-3)betaGal(1-4)betaGlcNAc), whereas the hemagglutinin displays higher affinity for a disialylated tetrasaccharide (alphaNeuAc(2-3)betaGal(1-3)[alphaNeuAc(2-6)]alphaG alNAc). The three-dimensional structure of the complex between M. amurensis leukoagglutinin and sialyllactose has been determined at 2.75-A resolution using x-ray crystallography. The carbohydrate binding site consists of a deep cleft that accommodates the three carbohydrate residues of the sialyllactose. The central galactose sits in the primary binding site in an orientation that has not been observed previously in other legume lectins. The carboxyl group of sialic acid establishes a salt bridge with a lysine side chain. The glucose residue is very efficiently docked between two tyrosine aromatic rings. The complex between M. amurensis hemagglutinin and a disialylated tetrasaccharide could be modeled from the leukoagglutinin/sialyllactose crystal structure. The substitution of one tyrosine by an alanine residue is responsible for the difference in fine specificity between the two isolectins. Comparison with other legume lectins indicates that oligosaccharide specificity within this family is achieved by the recycling of structural loops in different combinations. 相似文献
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A detailed study was made of the bark lectins of the legume tree Maackia amurensis using a combination of protein purification
and cDNA cloning. The lectins, which are the most abundant bark proteins, are a complex mixture of isoforms composed of two
types of subunits of 32 and 37 kDa, respectively. Isolation and characterization of the homotetrameric isoforms indicated
that the 32 kDa subunit exhibits a 100-fold stronger haemagglutinating activity than the 37 kDa subunit. Molecular cloning
confirmed that the two lectin subunits are encoded by different genes. The 32 kDa subunit is apparently encoded by a single
gene, whereas two highly homologous genes encode the 37 kDa subunit. A comparison of the deduced amino acid sequences of the
bark lectin cDNAs and the previously described cDNA encoding the seed haemagglutinin demonstrated that they are encoded by
different genes. Abbreviations: LECMAHb, cDNA clone encoding Maackia amurensis bark haemagglutinin; LECMALb, cDNA clone encoding
Maackia amurensis bark leucoagglutinin; MALb, Maackia amurensis bark leucoagglutinin; MAHb, Maackia amurensis bark haemagglutinin
This revised version was published online in November 2006 with corrections to the Cover Date. 相似文献
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Ronald L. Miller 《Journal of invertebrate pathology》1982,39(2):210-214
The slug, Limax flavus, contains a lectin that appears to be highly specific for sialic acid residues of glycoproteins. The carbohydrates which inhibited the hemagglutinating activity of the slug lectin and the concentration of the carbohydrate which gave a 50% inhibition are as follows: N-acetylneuraminic acid, 0.13 mm; N-glycolylneuraminic acid, 0.90 mm; d-glucosamine, 4.9 mm; d-galactosamine, 7.6 mm; N-acetyl-d-glucosamine, 23 mm; and N-acetyl-d-galactosamine, 24 mm. d-Galactose, d-glucose, d-mannose, α-methyl-d-glucoside, α-methyl-d-mannoside, l-arabinose, d-xylose, l-fucose, d-glucuronic acid, lactose, and sucrose were found to be ineffective as inhibitors of the hemagglutinating activity of the slug lectin. Hemagglutination by slug lectin was strongly inhibited by bovine submaxillary mucin and fetuin but not by sialic acid-free bovine submaxillary mucin or fetuin. 相似文献
6.
Purification of a sialic acid-specific lectin from the Indian scorpion Heterometrus granulomanus 总被引:1,自引:0,他引:1
A sialic acid-specific lectin, scorpin, has been purified to apparent homogeneity from the Indian scorpion Heterometrus granulomanus by affinity chromatography on equine submandibular gland glycopeptides linked to Sepharose and gel filtration on Sephadex G-200. The lectin has a molecular mass of 500 000 Da and was dissociated into single polypeptide chains of 15 000 Da, as determined by SDS gel electrophoresis in the presence of 2-mercaptoethanol. Scorpin is a glycoprotein containing 2.8% sugars. Its specificity was investigated by the inhibition of hemagglutination with various derivatives of sialic acid and other sugars. N-Acetylneuraminic acid gave better inhibition than N-glycoloylneuraminic acid but showed less inhibitory effect than sialyl-alpha(2----3)-lactose and disialyllactose. Among the sialoglycoconjugates tested, equine submandibular gland glycopeptide was found to be the most potent inhibitor. Scorpin showed a strong tendency to bind to carboxyl groups, since reduction of the carboxyl group of N-acetylneuraminic acid destroyed the inhibitory potency of this sugar. Furthermore, D-glucuronic acid inhibited hemagglutination whereas N-acetylglucosamine or N-acetylgalactosamine were not inhibitors. 相似文献
7.
A sialic acid-specific lectin was isolated from the albumin glands of the garden snail Cepaea hortensis by affinity chromatography on fetuin-Sepharose following gel filtration on Superdex 200. The purified native lectin showed a molecular mass of about 95 kDa by gel filtration and 100 kDa by SDS electrophoresis. It was cleaved by boiling in buffer containing SDS in three serological identical bands corresponding to molecular masses of about 24, 20 and 16 kDa, respectively. From these three fragments, only the 24- and the 20-kDa bands were found to be glycosylated. Only the three sugars mannose, galactose and N-acetylglucosamine could be detected in a molar ratio of 3:8.6:2. The oligosaccharide moieties seem to be N- and partially O-glycosidic bound. Isoelectric focusing (IEF) of the purified lectin revealed a heterogeneous pattern with bands in the pH range of 4.3-5.0. Isolated bands of different isoelectric points showed in SDS electrophoresis the same three fragments with molecular masses of 24, 20 or 16 kDa. The heterogeneity of the lectin was revealed either by IEF or amino acid sequencing of internal tryptic peptides. 相似文献
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An accurate, rapid, and sensitive method for characterizing the carbohydrate binding properties of lectins using a BIAcore apparatus and the detection method of surface plasmon resonance is described. As a model study, the sialic acid binding lectins from Sambucus nigra and Maackia amurensis, which are specific for the epitopes Neu5Ac(alpha2-6)Gal and Neu5Ac(alpha2-3)Gal, respectively, were chosen as suitable candidates. Two systems, one for the analysis of oligosaccharides and the other for glycoproteins, were developed after a rigorous analysis and evaluation of such parameters as binding conditions, buffers, and regeneration conditions. The systems take into account nonspecific binding, using the respective denatured lectin as negative blank, and avoid loss of activity: regeneration of the surface using either 10 mM NaOAc (pH 4.3) buffer (oligosaccharide system) or 20 mM HCl (glycoprotein system). The specificity of the lectins is well illustrated, while the kinetics parameters are shown to be sensitive to subtle changes in the recognized epitopes, and to be affected by steric hindrance. Surface plasmon resonance is a suitable technique for the analysis and characterization of lectins. 相似文献
10.
Interaction of the sialic acid-specific lectin carcinoscorpin with various sialoglycoproteins was studied by using radioiodinated lectin. The binding of carcinoscorpin was dependent not only on sialic acid content but also on the type of glycosidic linkage and form (branched or linear) of the carbohydrate chains. Carcinoscorpin has different classes of binding sites, and binding follows a phenomenon of positive co-operativity. The effect of Ca2+ concentration on the binding was studied, and the optimal concentration was found to be 0.02 M. Effect of pH, temperature and other bivalent metal ions are also reported. From haemagglutination- and precipitation-inhibition studies, it was concluded that carcinoscorpin has multispecificity towards acidic sugars, and its relation to the biological role of the lectin in the horseshoe crab is discussed. 相似文献
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A novel sialic acid-specific lectin from Phaseolus coccineus seeds with potent antineoplastic and antifungal activities 总被引:1,自引:0,他引:1
Jing Chen Bo Liu Na Ji Jing Zhou He-jiao Bian Chun-yang Li Fang Chen Jin-ku Bao 《Phytomedicine》2009,16(4):352-360
A novel lectin (PCL) with specificity towards sialic acid was purified from Phaseolus coccineus L. (P. multiflorus willd) seeds using ion exchange chromatography on CM and DEAE-Sepharose, and gel filtration on Sephacryl S-200 column. PCL was a homodimer consisting of 29,831.265 Da subunits as determined by gel filtration and MS. Also, PCL was a non-metaloprotein and its N-terminal 23-amino acid sequence, ATETSFSFQRLNLANLVLNKESS, was determined. Subsequently, MTT method, cell morphological analysis and LDH activity-based cytotoxicity assays demonstrated that PCL was highly cytotoxic to L929 cells and induced apoptosis in a dose-dependent manner. Using caspase inhibitors, a typical caspase-dependent pathway was confirmed. PCL also showed remarkable antifungal activity towards some plant pathogenic fungi. Furthermore, when sialic acid-specific activity was fully inhibited, cytotoxicity and antifungal activity were abruptly decreased, respectively, suggesting a significant correlation between sialic acid-specific site and its bi-functional bioactivities. 相似文献
13.
Kobayashi Y Kobayashi K Umehara K Dohra H Murata T Usui T Kawagishi H 《The Journal of biological chemistry》2004,279(51):53048-53055
A carbohydrate-binding protein was isolated from the carpophores of the mushrooms and designated the Chlorophyllum molybdites lectin (CML) based on its origin. The molecular mass of CML was 32 kDa, and it was composed of two 16-kDa monomers with no disulfide bonds. Monosaccharide analysis indicated that 12% of the mass of CML was carbohydrate and consisted of GlcNAc:GalNAc:Gal:Man:l-Fuc in a molar ratio of 1.5:1.9: 4.4:4.8:1.0. In the hemagglutination inhibition assay, CML exhibited the strongest binding specificity toward N-glycolylneuraminic acid (NeuGc) among the monosaccharides tested, whereas NeuAc did not inhibit the hemagglutination at all. GalNAc and Mealpha-GalNAc were also inhibitory at much higher concentrations than NeuGc. Among the glycoproteins, asialobovine submaxillary mucin (BSM) and porcine stomach mucin (PSM) showed strong inhibitory effects. In surface plasmon resonance analysis, asialo-BSM and PSM exhibited the strongest binding affinity. After co-injection of CML and NeuGc or GalNAc onto the asialo-BSM- or PSM-immobilized chip, the dissociation of CML from the immobilized PSM was accelerated by NeuGc and GalNAc, but the dissociation of CML from the immobilized asialo-BSM was only promoted by GalNAc. These results and the other surface plasmon resonance experiments allowed us to conclude that the binding of asialo-BSM to CML was because of an interaction between the lectin and the GalNAc residues of asialo-BSM, and both the NeuGc and GalNAc residues were responsible for the binding of PSM to CML. The results also suggested that CML had two different carbohydrate binding domains, one specific for NeuGc and the other for GalNAc. 相似文献
14.
The binding of fucose-containing glycoproteins by hepatic lectins. The binding specificity of the rat liver fucose lectin 总被引:2,自引:0,他引:2
The parameters that affect the interaction of ligands with a fucose-binding lectin from rat liver have been examined. 125I-Fucosyl-bovine serum albumin (Fuc-BSA) containing 50 residues of fucose/molecule was used as the standard ligand. At low initial concentrations of ligand (10 ng/ml) and lectin (140 ng/ml), the reaction reaches equilibrium at pH 7.8, 23 degrees C, within 40 min. The binding of ligands is Ca2+ dependent with half-maximal binding occurring at 54 microM Ca2+; of several metal ions tested, only Sr2+ partially replaced Ca2+. Binding was maximal between pH 7.6 and 8.6, fell slightly up to pH 10, but fell markedly below pH 7. The lectin-ligand complexes dissociated at low pH, on removal of Ca2+, or in the presence of a large excess of competing ligand. The apparent association constant (Ka) for Fuc-BSA was 1.75 X 10(8) M-1. The fucose content of the Fuc-BSA also influenced binding, with little apparent binding below 24 fucose residues/molecule and maximal binding from 40 to 50 fucose residues/molecule. With knowledge of the parameters influencing binding, sensitive reproducible assays for the lectin were developed. The binding specificity of the lectin was examined by measuring the inhibition of 125I-Fuc-BSA binding by neoglycoproteins, monosaccharides, and glycosides or by direct binding of neoglycoproteins. Galactosides and beta-linked fucosides were the best ligands among the neoglycoproteins, with much weaker binding by mannosyl- or N-acetylglucosaminyl-BSA. On the basis of the pattern of inhibition of Fuc-BSA binding by various monosaccharides and glycosides, it is possible to propose the conformations of saccharides that best fit the lectin-binding site. The C1 conformation of N-acetyl-D-galactosamine fits best, although other not obviously related monosaccharides such as L-fucose, L-arabinose, and D-mannose can also assume conformations that permit them to be effective inhibitors. The pattern of binding of neoglycoproteins to the lectin differs from that of other pure hepatic lectins. Thus, the fucose lectin has a high affinity for Fuc-BSA and galactosyl-BSA but a low affinity for N-acetylglucosaminyl-BSA. The galactose lectin binds only galactosyl-BSA and shows little binding with either N-acetylglucosaminyl-BSA or Fuc-BSA. In contrast, the mannose/N-acetylglucosamine lectin binds N-acetylglucosaminyl-BSA and Fuc-BSA but not galactosyl-BSA. 相似文献
15.
I J Goldstein 《Indian journal of biochemistry & biophysics》1990,27(6):368-369
The physiological role of plant lectins still remains a mystery. Nevertheless the carbohydrate binding properties of these proteins are a source of great fascination and have been applied to the study of glycoconjugates on cell surfaces and in solution. These matters are discussed. 相似文献
16.
A sialic acid-specific lectin from the mushroom Paecilomyces Japonica that exhibits hemagglutination activity and cytotoxicity 总被引:1,自引:0,他引:1
The mushroom Paecilomyces japonica, grown on the silkworm larvae, has been used in Asia as a nutraceutical, tea, and Chinese medicine. In the present study, a sialic acid-specific lectin has been purified from the mushroom P. japonica using affinity chromatography on a fetuin-agarose column. Electrophoretical analyses indicated that this lectin, designated P. japonica agglutinin (PJA), is an acidic protein with a molecular mass of 16 kDa, and has no intermolecular disulfide bonds. PJA induced hemagglutination activity in human ABO, mouse, rat, and rabbit erythrocytes. This activity was inhibited by sialic acid and sialoglycoproteins, but not by any other carbohydrates. PJA was stable at pH 4.0-8.0, and at temperatures below 55 degrees C. The activity of PJA was independent of EDTA and divalent cations. In addition, PJA exerts cytotoxic effects on the following cancer cell lines: human stomach cancer SNU-1, human pancreas cancer AsPc-1, and human breast cancer MDA-MB-231. 相似文献
17.
An unique specificity of a sialic acid binding lectin AchatininH, from the hemolymph of Achatina fulica snail 总被引:2,自引:0,他引:2
A sialic acid-binding lectin, AchatininH, from the hemolymph of Achatina fulica snail is found to be highly specific for 9-0-acetyl sialic acid. The binding specificity of AchatininH distinguishes it from other known sialic-acid specific lectins which usually show a broader range of specificity for sialic acid. It is even better than crab lectin which shows specificity for both 4- and 9-0-acetylated derivatives of sialic acid. This limited specificity of AchatininH appear to account for the fact that it agglutinates only rabbit, rat and guinea pig erythrocytes which contain 9-0-acetylated sialic acid but not horse (mainly contain 4-0-acetylated sialic acid), human, monkey, sheep, goat and chicken erythrocytes which contain either N-acetyl or N-glycolyl neuraminic acid but no 0-acetylated derivatives. This finding was further supported by the potent inhibition of hemagglutination by free 9-0-acetylated neuraminic acid and by several glyco shingolipids of human origin having 0-acetylated sialic acid. 相似文献
18.
Susann Teneberg Iréne Leonardsson Jonas Ångström Sarah Ehrlich-Rogozinski Nathan Sharon 《Glycoconjugate journal》1994,11(5):418-423
The specificity ofMoluccella laevis lectin was investigated by analysing its binding to glycosphingolipids separated on thin-layer chromatograms or adsorbed on microtitre wells. The binding activity of the lectin was highest for glycosphingolipids with terminal -linkedN-acetylgalactosamine, both in linear structures, as the Forssman glycosphingolipid, GalNAc3GalNAc3Gal4Gal4Glc1Cer, and in branched structures, as glycosphingolipids with the blood group A determinant, GalNAc3(Fuc2)Gal. In addition, the lectin bound, though considerably more weakly, to linear glycosphingolipids with terminal -linked galactose. When considering the use of theM. laevis lectin for biochemical and medical purposes this cross-reactivity may be of importance.
Nomenclature: The glycosphingolipid nomenclature follows the recommendations by the IUPAC-IUB Commission on Biochemical Nomenclature (CBN for Lipids:Eur J Biochem (1977)79:11–21,J Biol Chem (1982)257:3347–51, andJ Biol Chem (1987)262:13–18). It is assumed that Gal, Glc, GlcNAc, GalNAc, and NeuAc are of thed-configuration, Fuc of thel-configuration, and all sugars present in the pyranose form. 相似文献
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Mucin-specific lectin from Sambucus sieboldiana (SSA-M) reacts in Western blotting and ELISA with mucins from porcine stomach, bovine and ovine submaxillary glands, the human milk fat globule membrane, in vitro human ovarian, breast and colonic tumor cell lines, and mucins produced in vivo in the ascites of patients with endometrial and ovarian tumors, but not with fetal bovine fetuin or human transferrin. Sialidase treatment of these mucins led to an increase in the binding of SSA-M, suggesting that sialic acid is not part of the binding site for this lectin. Furthermore, sialic acid did not inhibit lectin binding. Treatment of asialomucin with O-glycanase decreased the binding of SSA-M, confirming the reactivity of the lectin with an O-linked carbohydrate. Treatment of mucins with trifluoromethanesulfonic acid, which removes all but core carbohydrate, led to an increase in the binding of SSA-M, suggesting that the lectin reacts with O-linked core glycans. Indeed, the increased reactivity after sialidase treatment of ovine submaxillary mucin suggests the lectin reacts with peptide-linked N-acetylgalactosamine (GalNAc), since more than 98% of the glycan chains attached to this mucin are sialylated GalNAc. The binding of SSA-M to sialidase-treated porcine mucin was inhibited strongly by GalNAc and disaccharides containing galactose (lactose, melibiose, and N-acetyllactosamine) but not by free galactose (Gal), suggesting that the glycan for optimum binding is Gal beta(1-3)GalNAc. This pattern of inhibition was different to other core glycan-reactive lectins tested, indicating that SSA-M is distinct, and should be of use in the isolation and characterisation of mucins and O-linked glycans. 相似文献
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Ueda H Matsumoto H Takahashi N Ogawa H 《The Journal of biological chemistry》2002,277(28):24916-24925
A lectin from the fruiting body of the Psathyrella velutina mushroom (PVL) was found to bind specifically to N-acetylneuraminic acid, as well as to GlcNAc (Ueda, H., Kojima, K., Saitoh, T., and Ogawa, H. (1999) FEBS Lett. 448, 75-80). In this study, the glycan sequences that PVL recognizes with high affinity on sialoglycoproteins were revealed. Among sialic acid-specific lectins only PVL could reveal the sialylated N-acetyllactosamine structure of glycoproteins in blotting studies, based on the dual specificity. The affinity of PVL to fetuin was measured by surface plasmon resonance to be 10(7) m(-1), which is an order of magnitude higher than those of Sambucus nigra agglutinin and Maackia amurensis mitogen, whereas affinity to asialofetuin was approximately 0 and to asialo-agalactofetuin was 10(8) m(-1), suggesting that PVL exhibits remarkably high affinities toward glycoproteins possessing trisialo- or GlcNAc-exposed glycans. Transferrin was separated into fractions that correspond to the sialylation states on an immobilized PVL column. Transferrin-possessing trisialoglycans containing alpha2,3-linked N-acetylneuraminic acid on the beta1,4-linked GlcNAc branch bound to the PVL column and eluted with GlcNAc; those containing only alpha2,6-linked sialic acids were retarded, whereas other transferrin fractions passed through the column. These results indicate that PVL is a lectin with potential for separation and detection of sialoglycoproteins because of its dual specificity toward sialoglycans and GlcNAc exposed glycans. 相似文献