Distribution of Bandeiraea simplicifolia lectin binding sites in the genital organs of female and male mice

Fluorescein isothiocyanate labelled type I lectin from Bandeiraea simplicifolia (BSA-I) known for its specific binding to alpha-D-galactopyranosyl and 2-acetamido-2-deoxy-D-galactose groups, has been used to map the distribution of the lectin specific binding sites in the genital organs of female and male mice. In non-pregnant female mice, strong lectin reactivity was restricted to the epithelium of the distal oviduct, the cervix and vagina. In pregnant mice strong BSA-I reactivity was also noted in the epithelium of uterine glands from the time of implantation on day 5 onward. In the testis BSA-I bound selectively to sperm but did not react with other cells in the seminiferous tubules. In the proximal caput epididymis BSA-I reacted with the epithelial cells, the underlying basement membranes and the intraluminal sperm. The intraluminal contents of the seminal vesicles reacted strongly with the lectin. Our data thus show a widespread but selective distribution of BSA-I lectin binding sites in the male and female genital organs and altered lectin binding in the uterus during pregnancy.     

Fluorescein-Conjugated Bandeiraea simplicifolia Lectin as a Marker of Endodermal, Yolk Sac, and Trophoblastic Differentiation in the Mouse Embryo

Abstract. The Bandeiraea simplicifolia lectin I (BSA-I) conjugated to fluorescein isothiocyanate was used as a histochemical reagent to study the mouse embryos from fertilization to early somitogenesis. No lectin binding could be detected on the embryonic cells in the preimplantation embryo. Lectin labeled intensely the zona pellucida. In the implanting embryos lectin binding was detected along the subtrophectodermal and Reichert's membrane, in the cytoplasm of the parietal and visceral endoderm, and the trophoblastic giant cells, but not in the ectodermal cells. Studies on explanted blastocyts cultured in vitro disclosed that the cytoplasmic BSA-I binding sites in trophoblastic cells develop gradually. In the 9-day somitic embryo BSA-I reacted with epithelial cells of the yolk sac, but not with the mesenchymal cells. A continuity between the lectin-reactive endoderm and the foregut epithelium could be demonstrated. These data indicated that BSA-I lectin can be used as a histochemical probe for endodermal (yolk sac) and trophoblastic differentiation in the peri-implantational mouse embryo.     

Griffonia simplicifolia lectins bind specifically to endothelial cells and some epithelial cells in mouse tissues

Summary The binding ofGriffonia simplicifolia agglutinin-I (GSA—I) and the isolectins GSA-I-AB₃ and GSA-I-B₄, having affinity for some -d-galactosyl andN-acetyl galactosaminyl residues was studied in different mouse tissues. In brain, cardiac muscle and skeletal muscle, the GSA-I-lectin conjugates showed prominent binding only to blood vessel endothelia. Similarly, in the liver and kidney cortex the GSA-I-conjugates selectively reacted with endothelial cells of the sinusoids and with intertubular and glomerular capillaries, respectively. However, a strong reactivity with the GSA-I-conjugates was additionally seen in the acinar cells of the pancreas, in the stratified squamous epithelia of skin and tongue, and in transitional epithelium. SDS—PAGE electrophoresis combined with the lectin-blotting technique indicated that a similar set of glycoproteins are responsible for the GSA-I binding, even in different tissues. Another lectin with specificity for -d-galactose, theMaclura pomifera agglutinin, displayed a distinctly different distribution of binding sites, mainly in the basement membranes, of all mouse tissues studied. The results suggest that some -d-galactosyl residues, recognized by the binding of GSA-I lectins, are preferentially expressed in endothelial cells of mouse tissues, and also provide further evidence that endothelial cells can present a highly specific surface glycosylation pattern.     

Alternative ultrastructural localization of Dolichos biflorus lectin binding sites in proton secreting parietal cells of mice

Summary High amount of N-acetyl-d-galactosamine specific lectin binding sites were detected on the canalicular membranes of human parietal cells. Our present model investigations on mice showed that the intracellular distribution of the terminal N-acetyl-d-galactosamine containing glycoprotein highly depends on the actual functional state of the parietal cells. In the normal gastric mucosa 40%–60% of parictal cells react positively after staining with horseradish peroxidase or biotin labelled Dolichos biflorus lectin. Ultrastructurally lectin binding sites occur mainly on the basolateral membrane infoldings in fed animals, while they are present exclusively on the canalicular membranes of fasting mice, suggesting that the alternative appearance of lectin binding sites on the opposite membrane areas of parietal cells is tightly coupled to their main function, to H⁺ secretion.     

Distribution of sugar residues in the bovine testis during postnatal ontogenesis demonstrated with lectin-horseradish peroxidase conjugates

Summary In the present study the distribution of various sugar residues in the cells of the male gonad during postnatal organogenesis was examined employing eight lectin-horseradish peroxidase conjugates (BS-I, ConA, DBA, PNA, RCA-I, SBA, UEA-I, WGA) on paraffin-embedded testicular tissue. The tissue was obtained from bull calves and young bulls of recorded age (4, 8, 16, 20, 25, 30, 40 and 52 weeks) and two adult bulls. During the whole observation period, lectin affinity in the developing testicular tubules was restricted to the germ cell line, while the Sertoli cells and their precursors remained completely unstained. DBA, a lectin with specific affinity to -d-GalNAc, served as a selective marker for prespermatogonia (PSG), the only precursors of bovine spermatogonia until the onset of spermatogenesis at week 30. -d-GalNAc, detected in the PSG Golgi zone and its vicinity, seems to play an important role during PSG proliferation and migration in the prepuberal testis. Concomitant with the differentiation of PSG into spermatogonia, the binding intensity of DBA to the Golgi zone of these cells decreased. After the gradual onset of spermatogenesis, the lectins revealed staining of Golgi complexes of most germ cell stages. Glycosylation of the cell components takes place in the Golgi complex, which explains the strong affinity of the lectins to this cell compartment. Inner and outer membrane of the acrosomal complex of spermatids, especially during Golgi and cap phase of spermiogenesis, were intensely stained with PNA, RCA-I and SBA. This staining disappeared in the maturation phase at the latest and indicates a role of terminal d-Gal-(13)-d-GalNAc, d-Gal and d-GalNAc during the formation of the sperm head and intraepithelial orientation of the spermatid. Other parts of the spermatid, such as the anulus and the cytoplasmic droplet, exhibited d-Gal, d-GlcNAc or sialic acid and d-GalNAc. In the intertubular tissue BS-I, RCA-I and UEA-I bound to vascular endothelia. Components of the intertubular extracellular matrix were stained with ConA (-d-Man), RCA-I (d-Gal), UEA-I (-l-Fuc) and WGA (d-GlcNAc or sialic acid).     

The monoclonal antibody,anti-asialoglycophorin from human erythrocytes,specific forβ-d-Gal-1-3-α-d-GalNAc-chains (Thomsen-Friedenreich receptors)

The monoclonal antibody 22.19 of IgM class obtained after immunization of BALB/c mice with asialoglycophorin of human erythrocyte membranes is described. The specificity of this antibody for -d-Gal-1-3--d-GalNAc- disaccharide chains (Thomsen-Friedenreich receptors) was established by studying its reactivity against various erythrocytes, glycoproteins and oligosaccharides and by comparison with two lectins, peanut agglutinin andVicia graminea lectin, which recognize these disaccharide chains.Abbreviations PNA peanut agglutinin - VgL Vicia graminea lectin - TF Thomsen-Friedenreich - HSA human serum albumin - MoAb monoclonal antibody     

Sex difference in the accumulation of d-β-aminoisobutyrate in organs of mouse after thymine loading

The concentration of d--aminoisobutyric acid (d-BAIB) in the liver and kidney was twice as high and dropped more slowly in the female mouse than in the male after an intraperitoneal injection of thymine. The concentration of -alanine, formed from uracil by the same enzyme system catalyzing formation of d-BAIB from thymine, was not different in the liver and kidney of both sexes after an intraperitoneal injection of uracil. After the intraperitoneal injection of d-BAIB, the concentration of BAIB in male liver decreased faster than that in female liver. Inhibition of d-BAIB: pyruvate aminotransferase caused by injection of d-cycloserine resulted in a significant increase in the concentration of BAIB in liver of both sexes after injection of thymine, but the concentration dropped more rapidly in the male. The activity of d-BAIB: pyruvate aminotransferase was not different in the livers of male and femalemice. Under the action of probenecid, an inhibitor of active transport systems, the sex difference in accumulation and disappearance of the amino acid in the liver was not observed. This suggested that the excretion of BAIB is more active in the renal tubules of the male mouse than in those of the female. However, the amount of BAIB excreted in the urine after injection of thymine was larger in the female mice than in the male mice. There may be another probenecid-sensitive enzyme for the disposal of BAIB in male mice.     

Histochemical study of Hurler's disease by the use of peroxidase-labelled lectins

Summary Peroxidase-labelled lectins specific for various carbohydrate residues were used as histochemical reagents in the investigation of Hurler's syndrome. Peanut lectin was used to detect terminald-galactose, wheatgerm lectin forN-acetyl-d-glucosamine, soybean lectin forN-acetyl-d-galactosamine,Tetragonolobus lotus lectin for -l-fucose andBandeiraea S. lectin for -d-galactose. It was found that Kupffer cells in the liver and splenic reticulo-endothelial cells contain acid mucopolysaccharides which bind lectins in paraffin sections after appropriate fixation. The pattern of lectin binding suggests that such cells contain significant amounts ofd-galactose,l-fucose,N-acetyl-d-galactosamine andN-acetyl-d-glucosamine. It is likely that the last named carbohydrate is present as a polymer. Neurones contain a different carbohydrate, rich in galactose and fucose but poor inN-acetyl-d-glucosamine. This compound is resistant to lipid extraction. Hepatocytes, as a rule, do not react with lectins, most likely because of loss of the more soluble mucopolysaccharides during fixation. The results are consistent with the biochemical data of Hurler's syndrome and indicate that lectins can be a useful tool for the investigation of the cytochemistry of storage disorders.     

The monosaccharide binding site of lentil lectin: an X-ray and molecular modelling study

The X-ray crystal structure of lentil lectin in complex with -d-glucopyranose has been determined by molecular replacement and refined to anR-value of 0.20 at 3.0 Å resolution. The glucose interacts with the protein in a manner similar to that found in the mannose complexes of concanavalin A, pea lectin and isolectin I fromLathyrus ochrus. The complex is stabilized by a network of hydrogen bonds involving the carbohydrate oxygens O6, O4, O3 and O5. In addition, the -d-glucopyranose residue makes van der Waals contacts with the protein, involving the phenyl ring of Phe123. The overall structure of lentil lectin, at this resolution, does not differ significantly from the highly refined structures of the uncomplexed lectin.Molecular docking studies were performed with mannose and its 2-O and 3-O-m-nitro-benzyl derivatives to explain their high affinity binding. The interactions of the modelled mannose with lentil lectin agree well with those observed experimentally for the protein-carbohydrate complex. The highly flexible Me-2-O-(m-nitro-benzyl)--d-mannopyranoside and Me-3-O-(m-nitro-benzyl)--d-mannopyranoside become conformationally restricted upon binding to lentil lectin. For best orientations of the two substrates in the combining site, the loss of entropy is accompanied by the formation of a strong hydrogen bond between the nitro group and one amino acid, Gly97 and Asn125, respectively, along with the establishment of van der Waals interactions between the benzyl group and the aromatic amino acids Tyr100 and Trp128.RL and FC are joint first authors.     

The presence of an endogenous lectin in early embryos ofXenopus laevis

Summary Xenopus laevis embryos were examined for the presence of endogenous carbohydrate binding proteins. Soluble extracts of cleavage, gastrula and neurula embryos are able to agglutinate trypsinized rabbit erythrocytes. Unlike other embryonic lectins this agglutination activity requires the presence of calcium ions but not of sulphydryl reducing agents. It is specifically inhibited by galactose and galactose containing derivatives. Thiodigalactoside is the most potent disaccharide inhibitor followed by lactose and melibiose respectively. Methyl -d-galactopyranoside is a more effective inhibitor than its anomer. N-acetyl-d-galactosamine, N-acetyl-d-glucosamine, methyl -d-mannopyranoside andl-fucose do not inhibit activity at concentrations at or above 25 mM. EDTA and EGTA are also strong inhibitors of this activity. -d-galactoside binding lectins present in the early chick embryo have been implicated in cell to cell and cell to substrate adhesiveness of extraembryonic chick endoderm cells. Since cells of the blastula inXenopus laevis possess surface receptors bearing terminal -d-galactoside groups it is possible that this -d-galactoside binding lectin may play a role in the control of cell surface mediated events during development.     

Properties of α-galactosidase II2 from Vicia faba seeds

-Galactosidase II² (MW 43 390) from resting Vicia faba L. seeds had been shown to possess d-glucose/d-mannose-specific lectin activity. Inhibition studies with monosaccharides and an examination of the effects of heat and pH on the catalytic and lectin activities of the enzyme indicate that the enzyme substrate and the lectin haptens bind at different sites on the protein. d-Mannosebinding has been investigated by equilibrium dialysis and spectrophotometrically. Both methods yield K_a values of approx. 3·10³ M^-1 for the interaction and there would appear to be two mannosebinding sites per molecule of enzyme protein. The lectin properties of V. faba -galactosidase II² have been discussed in relation to both V. faba lectin (favin) and other legume -galactosidases.Abbreviations con A concanavalin A - CM-cellulose carboxymethyl cellulose - MW molecular weight - PNPG p-nitrophenyl -d-galactoside - SDS sodium dodecyl sulphate - PAGE polyacrylamide-gel electrophoresis     

Differential binding characteristics and applications of dGalβ1→3dGalNAc specific lectins

Summary The binding properties of Arachis hypogaea (PNA), Bauhinia vurpurea alba (BPL), Maclura pomifera (MPL) and Sophora japonica (SJL) lectins were studied by quantitative precipitin and precipitin inhibition assays, demonstrating them to be most specific for dGal13dGalNAc residues. Additionally, each lectin had its own binding characteristic such as different binding abilities to dGal14dGlcNAc or dGal13dGlcNAc1linked oligosaccharides, and/or dGalNAc1linked to the Ser or Thr of the protein moiety. These differential binding characteristics can be used for investigating fine differences of the carbohydrate structure of the glycoconjugates, especially those having dGal13dGalNAc residues as terminal non-reducing ends.Abbreviations dGal d-galactopyranose - dMan d-mannopyranose - dGalNAc 2-acetamido-2-deoxy-d-galacto-pyranose - dGlcNAc 2-acetamino-2-deoxy-d-glucopyranose - LFuc L-fucose - NeuNAc N-acetylneuraminic acid - Ser serine - Thr Threonine - RCA Ricinus communis agglutinin - SBA Soy bean agglutinin (Glycine max) - HPA Helix pomatia agglutinin - DBL Dolichos biflorus lectin - GCL Geodia cydonium lectin     

Concanavalin A interactions with asparagine-linked glycopeptides. The mechanisms of binding of oligomannose,bisected hybrid,and complex type carbohydrates

The affinity of concanavalin A (Con A) for simple saccharides has been known for over 50 years. However, the specificity of binding of Con A with cell-surface related carbohydrates has only recently been examined in detail. Brewer and coworkers [J Biol Chem (1986) 261:7306–10; J Biol Chem (1987) 262:1288–93; J Biol Chem (1987) 262:1294–99] have recently studied the binding interactions of a series of oligomannose and bisected hybrid type glycopeptides and complex type glycopeptides and oligosaccharides with Con A. The relative affinities of the carbohydrates were determined using hemagglutination inhibition measurements, and their modes of binding to the lectin examined by nuclear magnetic relaxation dispersion (NMRD) spectroscopy and quantitative precipitation analyses. The equivalence zones (regions of maximum precipitation) of the precipitin curves of Con A and the carbohydrates indicate that certain oligomannose and bisected hybrid type glycopeptides are bivalent for lectin binding. From the NMRD and precipitation data, two protein binding sites on each glycopeptide have been identified and characterized. Certain bisected complex type oligosaccharides also bind and precipitate Con A, while the corresponding nonbisected analogs bind but do not precipitate the protein. The precipitation data indicate that the bisected complex type oligosaccharides are also bivalent for lectin binding, while the nonbisected analogs are univalent. The NMRD and precipitation data are consistent with different mechanisms of binding of nonbisected and bisected complex type carbohydrates to Con A, including different conformations of the bound saccharides.Abbreviations Con A Concanavalin A with unspecified metal ion content - CMPL Con A with Mn²⁺ and Ca²⁺ at the S1 and S2 sites respectively, in the locked conformation [12]; trisaccharide1, 3,6-di-O-(-d-mannopyranosyl)-d-mannose - -MDM methyl -d-mannopyranoside - NMRD nuclear magnetic relaxation dispersion, the magnetic field dependence of nuclear magnetic relaxation rates, in the present case, the longitudinal relaxation rate, 1/T₁, of solvent protons     

Lectin histochemistry of mouse vagina during the estrous cycle

Estrous cycle-related histochemical changes in the vaginal epithelium of sexually mature female mice were studied with 30 fluorescein isothiocyanate (FITC)-labeled lectins. On the basis of the staining pattern the lectins were divided into five groups: I, seventeen lectins that reacted with mucinous surface layer of proestrus. This group comprised two subgroups: Ia, seven lectins that reacted exclusively with the mucinous layer, and Ib, ten lectins that reacted with mucinous cells and the underlying squamous epithelium of proestrus; II, two lectins that reacted with squamous epithelium of proestrus only but were unreactive with mucinous cells; III, three lectins that reacted in a phase-specific manner with squamous epithelium; IV, six lectins that showed increased luminal surface reactivity in diestrus and/or metestrus; and V, eleven lectins that were unreactive with vaginal epithelium. These data indicate that the cyclic changes in the morphology of the vaginal epithelium are accompanied by distinct lectin reactivity patterns.     

Light and electron microscopical localization of concanavalin A lectin binding sites in rat epiphyseal chondrocytes

Summary Concanavalin A lectin binding sites have been detected within the cytoplasm of epiphyseal chondrocytes. Correlative light and electron microscopic results were obtained, indicating the presence of-d-mannose and/or -D-glucose residues detected by the lectin in the rough endoplasmic reticulum region. Quantitation of the electron microscopic cytochemical reaction also showed that the specific labelling was almost exclusively localized in the lumen of endoplasmic reticulum cisternae. No significant staining was found in other membrane compartments or extracellular matrix. This labelling pattern could be considered as the cytochemical evidence ofN-glycosylation processes occurring during the biosynthesis of cartilage extracellular matrix components by chondrocytes.     

Lectin binding by microvillous membranes and coated-pit regions of human syncytial trophoblast

Summary The distribution of saccharides on the microvillous membrane of the human syncytial trophoblast was investigated using ferritin conjugates of four lectins: concanavalin A (specific for -d-manno- and -d-glucopyranosyl residues), wheatgerm agglutinin (specific forN-acetylglucosamine),Limulus polyphemus lectin (specific forN-acetylneuraminic acid), andLotus tetragonolobus lectin (specific for -l-fucose). Concanavalin A and wheatgerm agglutinin (WGA) reacted strongly with the surface membrane and ferritin deposits were also observed in coated pit regions of the membrane. Lectins fromL. polyphemus andL. tetragonolobus, however, reacted only weakly with the microvillous border and neither reacted with coated pits.Enhanced agglutinability of trophoblast cells in comparison with other foetal cells from the same conceptus was seen with WGA. This agglutination was inhibited by addition of acetylglucosamine or by a solubilized membrane fraction which was bound by a column of WGA-Sepharose. The membrane fraction which did not bind to the column did not inhibit agglutination. Electrophoresis of the WGA-bound membrane proteins revealed six subunits, the major band having an apparent mol. wt. of 55 000. A protein of this mol. wt was also seen in coated vesicles isolated from equivalent human placentae.     

Location of amino acid and carbohydrate transport sites in the surface membrane of normal and transformed mammalian cells

Summary Amino acid and carbohydrate transport in normal and malignant transformed hamster cells was studied after binding of the protein Concanavalin A (Con. A) to the surface membrane. Experimental conditions were used so that a similar number of Con. A molecules were bound to both types of cells. The transport of amino acids was inhibited after Con. A binding in the transformed cells but not in normal cells. This was found with the metabolizable amino acidsl-leucine,l-arginine,l-glutamic acid, andl-glutamine, and with the non-metabolizable amino acids cycloleucine and -aminoisobutyric acid. Transport ofd-glucose andd-galactose was more inhibited by Con. A in transformed than in normal cells, and in both types of cellsd-glucose was inhibited more thand-galactose. The inhibition by Con. A on transport was specific, since there was no effect on the transport ofl-fucose in either normal or transformed cells. Con. A also did not effect the entry of 3-0-methyl-d-glucose.These observations can be used to locate amino acid and carbohydrate transport sites in the surface membrane in relation to the binding sites for Con. A. The results indicate that Con. A sites are associated in normal cells with transport sites ford-glucose and to a lesser extentd-galactose, and in transformed cells with transport sites for amino acids and to a greater extent than in normal cells withd-glucose andd-galactose. Malignant transformation of normal cells therefore results in a change in the location of amino acid and carbohydrate transport sites in the surface membrane in relation to the binding sites for Con. A.     

Binding sites of Ulex europaeus-lectin I in human parotid gland

Summary Twenty non-neoplastic parotid glands (removed during neck dissection for regional tumours) were examined for cellular and subcellular binding sites of Ulex europaeus-lectin I (UEA-I), a lectin reported to be specific for -L-fucose. For light microscopy, an extended peroxidase-antiperoxidase method was applied; for the evaluation of the subcellular localization of bound lectin, three of these glands were examined following immunocryoultramicrotomy and staining by the protein A-gold technique.In addition to the known cytoplasmic affinity of UEA-I for capillary endothelium, acinar cells bound the lectin within the cytoplasmic compartment; the number and distribution of stained acinar cells varied among individuals. Furthermore, cytomembrane-bound labelling that occurred most markedly at the luminar surface was observed in striated-duct epithelium.Using the electron microscope, protein A-gold particles were seen in zymogen granules and in Golgi cisternae of serous acinar cells; primary saliva secreted in the lumina exhibited strong labelling; serous acinar cells had binding sites on their cell membranes, striated-duct epithelium had binding sites on its surface membrane and in the vicinity of apical vesicles. Our results show that UEA-I is a useful tool for the study of the structure and functional states of the parotid gland epithelium and its associated pathological alterations.Dedicated to Prof. Dr. med. G. Seifert on the occasion of his 65th birthday     

Synthesis ofN-Acetylglucosamine derivatives as probes for specificity of chicken hepatic lectin

Syntheses of the following compounds are described: 6-(Trifluoroacetylamino)hexyl 2-acetamido-2,6-dideoxy--d-glucopyranoside and 2-acetamido-2-deoxy--d-xylopyranoside, two allyl 2-acetamido-2-deoxy--d-glucopyranosiduronic acid derivatives, and several allyl 2-acylamido-2-deoxy--d-glucopyranosides having different acyl groups. These and other compounds were used as inhibitors in the binding assay for the chicken hepatic lectin specific forN-acetylglucosamine. We found that: 1) The inhibitory potency ofN-acylglucosamine derivatives decreased progressively with increase in the size of acyl group, 2) absence of either 3-or 4-OH group ofN-acetylglucosamine lowered the binding affinity more than 100-fold, and 3) the presence of a negatively charged group (carboxylic acid) at the C-6 position did not lower the affinity. The first two items are similar to the mammalian hepatic galactose/N-acetylgalactosamine lectins, but the last item is in a strong contrast to the mammalian lectins.Abbreviations XyLNAc N-acetyl-d-xylosamine - BSA bovine serum albumin - NeuAc N-acetylneuraminic acid - GlcNAc34-BSA amidino-type neoglycoprotein [6] containing on the average 34N-acetylglucosaminyl residues per BSA molecule     

Characterization of carbohydrate combining sites of Bryohealin,an algal lectin from <Emphasis Type="Italic">Bryopsis plumosa</Emphasis>

Bryohealin is a lectin involved in the wound-healing process of the marine green alga Bryopsis plumosa. In the previous purification study, it has been shown that lectin was composed of two identical subunits of 27 kDa, cross-linked by disulfide bond, and showed binding specificity to N-acetyl-d-glucosamine and N-acetyl-d-galactosamine (GlcNAc and GalNAc, respectively). To determine if the lectin recognize the two different sugars at the same binding domain, the carbohydrate binding sites of Bryohealin was analyzed using chromatography and chemical modification methods. Results showed that the same binding site of the lectin was responsible for the recognition of two sugars, GalNAc as well as GlcNAc. Chemical modification studies showed that hemagglutinating activities of Bryohealin were not affected by modification of histidine, tryptophan, aspartic acid, and glutamic acid. When arginine residues were modified with 1,2-cyclohexanedione, the activity of Bryohealin rapidly decreased. The sugar binding sites remained intact when the lectin was treated with inhibitory sugars (0.2 M GalNAc and/or GlcNAc) prior to 1,2-cyclohexanedione treatment. The sugar binding domain of Bryohealin was predicted from the MALDI-TOF analysis and the full cDNA sequence of the lectin gene.