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PubMed1991

Lectin histochemistry of trachea and lung of healthy turkeys (Meleagris gallopavo) and turkeys with pneumonia.

Thirteen lectins were used to characterize lectin-binding specificity of glycoconjugates on sections of formalin-fixed lung and trachea from seven normal turkeys, two turkeys with acute pneumonia, and two turkeys with chronic pneumonia. Neuraminidase was used to digest sialic acid residues. One N-acetylgalactosamine-binding lectin and two N-acetylgalactosamine/galactose-binding lectins stained the apical membrane and cytoplasm of multifocal cells that lined air atria and hyperplastic granular cells. Other lectins in these groups stained ciliated cells of the trachea and bronchi and air capillary epithelial cells. Sialic acid residues were on apical surfaces of ciliated and nonciliated tracheal and bronchial lining cells, air capillary epithelial cells, and vascular endothelial cells. Mannose/glucose-binding lectins stained reticular and elastic fibers in the lamina propria of trachea, primary and secondary bronchi, and the tunica adventitia of arteries and veins. By transmission electron microscopy, colloidal gold-Arachis hypogaea (peanut agglutinin) labeled microvilli on the apical surface of mature granular cells. The L-fucose-binding lectin, in addition to several other lectins, stained nonspecifically in both trachea and lung. These studies show that granular cells that line air atria can be identified with lectins of N-acetylglucosamine and N-acetylgalactosamine/galactose groups, and that apical surfaces of epithelial cells and endothelial cells in the trachea and lung express terminal sialic acid residues.

Acute Disease↗
PubMed2005

Pseudomonas aeruginosa lectins I and II and their interaction with human airway cilia.

The bacterium Pseudomonas aeruginosa (PA) produces two carbohydrate binding lectins, designated PA lectin-I and lectin-II (PA-IL, PA-IIL). Both lectins are used by the bacterium to adhere to the glycocalyx of mammalian cells. In addition, the lectins immobilize ciliary beat. The kinetics of ciliary beat inhibition by each individual lectin have been analysed; however, their joint action on cilia has not been reported. Here we demonstrate that PA-IL and PA-IIL inhibit ciliary beat in a similar time-dependent manner. If applied simultaneously, ciliary beat inhibition after five hours of incubation was weaker than if each lectin was applied separately. Thus it can be hypothesized that the lectins compete for the same binding site(s) of the glycocalyx. Sugar inhibition experiments demonstrate that D-galactose and L-fucose inhibit both lectins, although clear preferences of D-galactose for PA-IL and of L-fucose for PA-IIL exist. These interactions have to be kept in mind when designing sugar-based therapies.

Adhesins, Bacterial↗
PubMed1997

Lectin histochemistry in rat liver fibrosis induced by heterologous serum sensitization.

The localization of carbohydrates in rat livers with fibrosis induced by heterologous serum was examined by lectin histochemical and biochemical techniques. Twenty-four lectins were used to visualize the different carbohydrates in paraffin sections of normal and fibrotic liver tissues. No differences in staining patterns of these lectins were observed between normal and fibrotic livers in hepatocyte cell membranes including bile canaliculi, sinusoidal endothelial, or bile ductal cells. Kupffer cells strongly stained with Vicia villosa agglutinin (VVA) were seen only in the periportal zone of the normal liver, but they were observed in the periportal zone and scattered throughout the pseudolobular zone in the fibrotic liver. The cytoplasm of some hepatocytes was strongly stained by Bandeiraea simplicifolia lectin-I (BSL-I). BSL-I positive hepatocytes in normal liver were localized in the periportal zone, but those in the fibrotic liver were scattered in the periportal and perifibrous zones. After polyacrylamide gel electrophoresis of liver glycoproteins, differences in molecular sizes of BSL-I positive glycoproteins (79 and 81 kD) were detected by lectin blotting. Cell density of perifibrous BSL-I positive hepatocytes may be useful as a diagnostic parameter for liver fibrosis and/or cirrhosis. Two distinct staining patterns with twelve lectins were observed in fibrotic septa of the fibrotic liver. The fibrotic septa were stained with six characteristic lectins, and the centrilobular septa were stained with all these twelve of lectins. Histopathological assessment of the centrilobular fibrotic septa stained with these characteristic lectins may contribute to the diagnosis and prognosis of hepatic fibrosis.

Animals↗
PubMed1983

Generation of fibronectin receptors on macrophages by wheat germ lectin.

A chymotrypsin-derived and 125I-labelled 125-kDa fragment of human plasma fibronectin which contained the cell binding site, was only weakly bound by peritoneal macrophages of guinea pigs and binding was not saturable. In presence of wheat germ lectin binding increased proportionally to the logarithm of the lectin concentration. Association of 125I-fragment with cells was partially prevented by non-labelled fragment indicating a saturable receptor-ligand interaction. An apparent affinity constant of about 2--4 x 10(-5) M was evaluated. A considerable fraction of the cell-bound 125I-fragment resisted removal by proteases suggesting that it was internalized. In order to investigate an influence of wheat germ lectin on the binding of 125I-fibronectin by the cells the macrophages were preincubated with the lectin followed by washing and evaluation of 125I-fibronectin binding. A simultaneous incubation of the cells with 125I-fibronectin and lectin was impractical due to partial interaction of the two proteins giving rise to some unspecific precipitates. Preincubation with wheat germ lectin considerably improved the capacity of the macrophages for binding of 125I-fibronectin. Again the binding of 125I-labelled protein could be restricted by unlabelled one. N-acetyl-glucosamine inhibited the binding of 125I-fibronectin by wheat germ lectin-treated cells if applied in the preincubation phase and more effectively, if applied in the final 125I-fibronectin binding assay. N-Acetylneuraminic acid also inhibited this step. In addition to wheat germ lectin concanavalin A was capable of generating fibronectin receptors on the cell surface. Soy bean lectin, however, was ineffective.

Animals↗
PubMed2005

Differential activity of a lectin from Solieria filiformis against human pathogenic bacteria.

A lectin isolated from the red alga Solieria filiformis was evaluated for its effect on the growth of 8 gram-negative and 3 gram-positive bacteria cultivated in liquid medium (three independent experiments/bacterium). The lectin (500 microg/mL) stimulated the growth of the gram-positive species Bacillus cereus and inhibited the growth of the gram-negative species Serratia marcescens, Salmonella typhi, Klebsiella pneumoniae, Enterobacter aerogenes, Proteus sp, and Pseudomonas aeruginosa at 1000 microg/mL but the lectin (10-1000 microg/mL) had no effect on the growth of the gram-positive bacteria Staphylococcus aureus and B. subtilis, or on the gram-negative bacteria Escherichia coli and Salmonella typhimurium. The purified lectin significantly reduced the cell density of gram-negative bacteria, although no changes in growth phases (log, exponential and of decline) were observed. It is possible that the interaction of S. filiformis lectin with the cell surface receptors of gram-negative bacteria promotes alterations in the flow of nutrients, which would explain the bacteriostatic effect. Growth stimulation of the gram-positive bacterium B. cereus was more marked in the presence of the lectin at a concentration of 1000 microg/mL. The stimulation of the growth of B. cereus was not observed when the lectin was previously incubated with mannan (125 microg/mL), its hapten. Thus, we suggest the involvement of the binding site of the lectin in this effect. The present study reports the first data on the inhibition and stimulation of pathogenic bacterial cells by marine alga lectins.

Anti-Bacterial Agents↗
PubMed1985

Eimeria meleagrimitis sporozoites: effect of lectins on invasion of cultured cells.

Exposure of primary turkey kidney cell cultures to 100 micrograms/ml wheat germ agglutinin (WGA), a lectin that binds N-acetylglucosamine (NAcGl), significantly inhibited invasion of the cells by Eimeria meleagrimitis sporozoites. However, neither succinyl-WGA, a lectin that retains an affinity for NAcGl but does not bind sialic acid, nor pokeweed mitogen, another lectin that binds NAcGl, similarly inhibited invasion. Collectively, the data suggested that the inhibition of invasion may be caused primarily by binding of WGA to anionic moieties on the host cell surface and not to specific binding to NAcGl. Exposure of cells to concanavalin A, phytohemagglutinin, or Ricinus communis lectin, as well as with pokeweed mitogen, failed to inhibit invasion by the sporozoites. Ultraviolet microscopy, using fluorescein-conjugated lectins, showed that the lectins had bound to the surface of the cultured cells. No binding of NAcGl or lectins to the surfaces of the sporozoites was demonstrated by either the clumping of the parasites in different lectin concentrations or the use of fluorescein-conjugated lectins. However, exposure of E. meleagrimitis sporozoites to NAcGl increased invasion of untreated cells by 50%.

Animals↗
PubMed1984

Lectins discriminate between pathogenic and nonpathogenic South American trypanosomes.

Cell surface carbohydrates of Trypanosoma cruzi, Trypanosoma rangeli, and Trypanosoma conorhini were analyzed by a micro-agglutination assay employing 27 highly purified lectins and by binding assays using various 125I-labeled lectins. The following seven lectins discriminated between the trypanosomes: 1) tomato lectin (an N-acetyl-D-glucosamine-binding protein), both in purified form and as crude tomato juice; 2) Bauhinea purpurea and Sophora japonica lectins (both N-acetyl-D-galactosamine-binding proteins), which selectively agglutinated T. cruzi; 3) Vicia villosa (an N-acetyl-D-galactosamine-binding protein) which was specific for T. rangeli; 4) peanut lectin (a D-galactose-binding protein) both in purified form and as crude saline extract; and 5) Ulex europaeus and Lotus tetragonolobus (both L-fucose-binding proteins) lectins which reacted only with T. conorhini. Binding studies with 125I-labeled lectins were performed to find whether unagglutinated cells of the three different species of trypanosomes might have receptors for these lectins, in which case absence of agglutination could be due to a peculiar arrangement of the receptors. These assays essentially confirmed the agglutination experiments.

Agglutination Tests↗
PubMed1998

Serum IgM antibody response to the galactose-inhibitable adherence lectin of Entameoba histolytica.

An ELISA for detection of serum IgM antibodies to the galactose-inhibitable adherence lectin of Entamoeba histolytica revealed that 2.8% of uninfected controls, 0.0% of controls infected with other parasites, 13.4% of asymptomatic amebic infections, 55% of colitis patients, and 77% of amebic liver abscess patients from Cairo, Egypt and Durban, South Africa had serum anti-lectin IgM antibodies. Of acute amebic colitis patients with symptoms for less than one week, only 6% possessed serum IgG anti-lectin antibodies, yet 45% had serum IgM antibodies to the amebic lectin. This compares with 65% of sera in acute colitis patients positive for lectin antigen as determined by ELISA with anti-lectin monoclonal antibodies. In conclusion, an ELISA for serum anti-lectin IgM antibodies appears to have greater clinical utility in the setting of acute amebic colitis than an ELISA for anti-lectin IgG antibodies, but is no more sensitive than an ELISA for detection of lectin antigen in sera.

Animals↗
PubMed1999

Sialic acid-specific lectin-mediated adhesion of Tritrichomonas foetus and Tritrichomonas mobilensis.

Tritrichomonas foetus is an obligate parasite of the bovine urogenital tract producing infection associated with inflammatory changes, abortion, and infertility, Tritrichomonas mobilensis was isolated from squirrel monkey colon, and symptoms involve diarrheal complications. Both tritrichomonads produced hemagglutinins with the properties of sialic acid-specific lectins. Assays on the adherence of these protozoans to Chinese hamster ovary (CHO) cells and to bovine cervical and monkey colon mucus were performed to assess the function of the lectins in adhesion. Sialic acid at concentration as low as 2 mM inhibited the adhesion to CHO cells, less effectively to the mucus. Predigestion with Clostridium perfringens sialidase prevented the adhesion to both epithelial cells and the mucus. Inhibition of endogenous sialidases with 2,3-dehydro-2-deoxy-NeuAc increased the adhesion of T. mobilensis to CHO cells. Specific anti-T. foetus lectin (TFL) and anti-T. mobilensis lectin (TML) antibodies inhibited adhesion of the trichomonads to the epithelial cells and to the mucus. TFL histochemistry disclosed the presence of lectin ligands on keratinized vaginal epithelia, cervical mucosa, and mucin and on endometrial glands and their secretions. TML histochemistry showed reactivity with the luminal membranes of colonic glandular epithelium and less with the colonic mucin. Both lectins bound to the surface membrane of CHO cells. Anti-lectin antibodies showed granular cytoplasmic and strong membrane localization of the lectins in both tritrichomonads. Although the 2 tritrichomonads have different habitats, the results indicate that both these protozoa use lectins with sialic acid specificity for adhesion to mucosal surfaces.

Animals↗
PubMed1999

Generation of recombinant fragments of CD11b expressing the functional beta-glucan-binding lectin site of CR3 (CD11b/CD18).

CR3 (Mac-1; alphaMbeta2 integrin) functions as both a receptor for the opsonic iC3b fragment of C3 triggering phagocytosis or cytotoxicity and an adhesion molecule mediating leukocyte diapedesis. Recent reports have suggested that a CR3 lectin site may be required for both cytotoxic responses and adhesion. Cytotoxic responses require dual recognition of iC3b via the I domain of CD11b and specific microbial surface polysaccharides (e.g., beta-glucan) via a separate lectin site. Likewise, adhesion requires a lectin-dependent membrane complex between CR3 and CD87. To characterize the lectin site further, a recombinant baculovirus (rBv) system was developed that allowed high level expression of rCD11b on membranes and in the cytoplasm of Sf21 insect cells. Six rBv were generated that contained truncated cDNA encoding various CD11b domains. Immunoblotting of rBv-infected Sf21 cells showed that some native epitopes were expressed by five of six rCD11b fragments. Lectin activity of rCD11b proteins was evaluated by both flow cytometry with beta-glucan-FITC and radioactive binding assays with [125I]beta-glucan. Sf21 cells expressing rCD11b that included the C-terminal region, with or without the I-domain, exhibited lectin activity that was inhibited by unlabeled beta-glucan or anti-CR3 mAbs. The smallest rCD11b fragment exhibiting lectin activity included the C-terminus and part of the divalent cation binding region. The beta-glucan binding affinities of the three C-terminal region-containing rCD11bs expressed on Sf21 cell membranes were not significantly different from each other and were similar to that of neutrophil CR3. These data suggest that the lectin site may be located entirely within CD11b, although lectin site-dependent signaling through CD18 probably occurs with the heterodimer.

Animals↗
PubMed2000

Binding of some Diocleinae lectins to the mucosal surfaces of the eye and mouth.

Controlled delivery of diagnostic or therapeutically active agents to the eye and oral cavity is limited by the efficient removal mechanisms that exist in these areas. In this work, recently isolated, purified and characterised Brazilian bean lectins were examined for their ability to bind to the buccal and sublingual mucosa, cornea and conjunctiva of rat, with regard to their potential therapeutic use. These lectins were labelled using biotin (long arm) N-hydroxysuccinimide ester and a technique developed at the University of Portsmouth. Evaluation of binding after 15 min contact with intact (unfixed) rat eye and oral cavity epithelia was completed using a standard histological procedure involving the avidin-biotin reaction and formation of a brown precipitate with a 3,3'-diaminobenzidine solution. When examined by affinity chromatography, all lectins studied were found to show high percentage biotinylation (i.e. less than 4% by weight of total lectin was eluted from the column). The lectin:biotin ratio was estimated using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE), and ranged between 1:2 (Cratylia floribunda) and 1:10.4 (Canavalia brasilensis). As with previous studies, all lectins bound to the mucosal surfaces to some extent--the Cratylia floribunda lectin appearing to be the most promising, whereas the Dioclea violacea lectin showed comparatively weak binding. In all cases, the presence of glucose (the hapten sugar) reduced or eliminated binding. It was concluded that glucose-mannose binding sites are present on these surfaces, and the lectins from C. floribunda showed a particular affinity for these.

Animals↗
PubMed2004

Approach to the comprehensive analysis of glycoproteins isolated from human serum using a multi-lectin affinity column.

Glycosylation is one the most common post-translational modifications (PTM) and glycoproteins play fundamental roles in a diversity of biological processes. The development of an analytical approach to the study of variation of glycosylation patterns in serum samples has been hindered by the structural heterogeneity of this post-translational modification and the complexity of serum proteome. We have used the ability of different lectins to recognize specific glycosylation motifs to develop a specific affinity system that can achieve a comprehensive capture of serum glycoproteins. In a preliminary investigation, we evaluated the ability of five commonly used immobilized lectins to capture glycoproteins from human serum. SDS-PAGE analysis showed each lectin was able to enrich a subset of the serum glycoproteome and overlaps in lectin specificity were indeed observed. Based on these results and with the goal of studying the extent of the human serum glycoproteome, we then developed a multi-lectin affinity column containing Concanavalin A (Con A), Wheat germ and Jacalin lectin. The selection of lectins was also based on the known N-linked and O-linked glycan structures that are considered representative of the serum proteome. We then demonstrated that the capture of glycoproteins was specific, efficient and reproducible with this multi-lectin column. The results obtained with this affinity step indicated that about 10% of human serum proteins are glycosylated (weight/weight) and, after removal of six high abundance proteins, including albumin, at least 50% of the remaining proteins were glycosylated. We then evaluated the use of this affinity column to monitor changes in the pattern of glycosylation in serum samples by a controlled, stepwise release of the captured proteins from the multi-lectin affinity column with specific displacers.

Chromatography, Affinity↗
PubMed1992

Subunit molecular mass assignment of 14,654 Da to the soluble beta-galactoside-binding lectin from bovine heart muscle and demonstration of intramolecular disulfide bonding associated with oxidative inactivation.

The soluble dimeric beta-galactoside-binding lectin (subunit molecular mass, approximately 14 kDa) of bovine heart muscle, in common with the 14-kDa lectins of several other animal species, displays carbohydrate-binding activity when it is in the reduced state, but the purified lectin loses this activity upon oxidation. In the present study, the presence of any post-translational modification and the mechanism of the oxidative inactivation have been investigated by analyses of the reduced and oxidized forms of the purified bovine lectin by electrospray ionization-mass spectrometry (ESI-MS) and by liquid secondary ion mass spectrometry (LSIMS) of tryptic and peptic peptides. By ESI-MS, the molecular mass of the reduced lectin is determined to be 14,654.6 +/- 0.9 Da, and that of the oxidized lectin is 14,649.3 +/- 1.1 Da. These masses correspond to the amino acid sequence of the protein with the cysteines having free sulfhydryl groups in the reduced state and forming disulfide bonds in the oxidized state. There is no evidence of post-translational modification in either lectin form except for monoacetylation already predicted for alanine at the blocked N-terminal end. Pronounced differences in charge distribution in the electrospray ionization mass spectra of the reduced and oxidized lectin, reflecting a change in the number of accessible protonation sites in the oxidized protein, are consistent with the protein being held in an altered conformation by covalent bonding. The results of LSIMS analyses of tryptic and peptic peptides in conjunction with Edman sequencing indicate that disulfide bonding occurs predominantly between Cys2 and Cys130, Cys16 and Cys88, and Cys42 and Cys60. There is no evidence of oxidation of Trp68. These results, taken together with observations that almost the complete polypeptide chain is necessary for the functional integrity of the carbohydrate recognition domain (Abbott, W. M., and Feizi, T. (1991) J. Biol. Chem. 266, 5552-5557) point to intramolecular disulfide bonding with a change in protein folding and conformation as the mechanism of oxidative inactivation of the purified bovine lectin.

Amino Acid Sequence↗
PubMed1988

Differential binding properties of Ga1NAc and/or Ga1 specific lectins.

Grouping of lectin binding properties, based on determinant structure rather than monosaccharide inhibition pattern, should facilitate the selection of lectins as structural probes for glycans as well as for the interpretation of the distribution and the properties of the carbohydrate chains on the cell surface. Based on the binding specificities studied with glycan by precipitin-inhibition, competitive-binding and hemagglutinin-inhibition assays, twenty Ga1 and/or Ga1NAc specific lectins have been divided into six classes according to their specificity for the disaccharide as all or part of the determinants and Ga1NAc alpha 1----Ser(Thr) of the peptide chain. The differential affinities of these lectins were characterized by quantitative precipitin assay. Abbreviation of the following six lectin determinants can also be used to classify these lectins. (1) F determinant (GalNAc alpha 1----3GalNAc, Forssman specific disaccharide). (2) A (Af) determinant (GalNAc alpha 1----3Gal, Human blood group A specific disaccharide; Af, fucosylated A, (GalNAc alpha 1----3 [LFuc alpha 1----2]Gal). (3) Tn determinant (GalNAc alpha 1----0 to Ser (Thr) of the protein core, Tn antigen). (4) T determinant (T antigen, Gal beta 1----3GalNAc alpha 1----0 to Ser (Thr) of the protein core, the mucin type sugar sequence on the human erythrocyte membrane or Gal beta 1----3GalNAc beta 1---- at the nonreducing end of ganglioside). (5) I and II determinants (human blood group type I and II carbohydrate sequences). Most of the lectins reactive to Gal beta 1----4GlcNAc (II) are also reactive to Gal beta 1----3GlcNAc (I). Lectin I (II) determinants (i.e. Gal beta 1----3 (4) GlcNAc residues) can be found at the nonreducing end of the carbohydrate chains derived from either N-glycosidic or O-glycosidic linkages. (6) B determinant (Gal alpha 1----3Gal, Human blood group B specific disaccharide). Their carbohydrate specificities are classified as following: (Table see text). The differential binding properties of lectins can be defined from comparisons of their carbohydrate specificities listed above.

Acetylgalactosamine↗
PubMed1988

[Localization of glycoconjugates in tissues of mammals revealed by means of labeled lectins].

Histochemical peculiarities on binding of castor-oil plant, soybean and lentil lectins with tissues of the mucous membrane in the stomach, small and large intestine have been studied in the human being, rat, mouse, as well as the lectins mentioned and the maize agglutinin with the nervous tissue of the rat cerebral tissue. The reactions are carried out with nonfixed cryostat and deparaffinized histological slices. Lectins labelled with horseradish peroxidase are used. Certain specific peculiarities and differences concerning the lectin binding with tissues of the organs studied are determined. Predominant binding is noted of the soybean lectin with parietal and mucin-producing cells of the stomach, with epitheliocytes of the duodenal glands, with the brush border of the epithelial cells of the intestinal villi. The lentil and castor-oil plant lectins make contours of the basal membrane epithelium in the stomach and intestine. The lentil lectin also reacts with the germinative centers of the stomach lymphatic nodules and the castor-oil plant agglutinin--with the brush border of the small intestine epitheliocytes. The lectins used are predominantly bound with neurons of the subcortical formations of the rat brain and cerebral cortex. By means of labelled lectins of lentil, soybean, and castor-oil plant it is possible to reveal certain modifications of the rat small intestine glycoconjugates produced by means of the immortelle extract.

Animals↗
PubMed1986

The isolation of a rat alveolar macrophage lectin.

A lectin in rat alveolar macrophage membranes with a high affinity for binding ligands containing L-fucose and N-acetyl-D-glucosamine has been isolated by affinity chromatography on Fuc-BSA-Sepharose (where Fuc is fucosyl and BSA is bovine serum albumin). The lectin was extracted from rat lung homogenates with Triton X-100, absorbed from the extract onto Fuc-BSA-Sepharose in the presence of Ca2+ and eluted by removal of Ca2+. After a second adsorption to and elution from Fuc-BSA-Sepharose, three protein species were detected electrophoretically in fractions that bind Fuc-BSA. One, which was the mannose/N-acetylglucosamine lectin (Mr = 32,000) found earlier in hepatocytes, was removed by adsorption on anti-lectin IgG-Sepharose. Another (Mr = 46,000) was removed by adsorption to Fuc-BSA-Sepharose and elution with galactose. The remaining lectin (Mr = 180,000) bound fucose and N-acetylglucosamine but not galactose. Binding was maximal between pH 6.5 and 9.0 and dependent on Ca2+. Immunocytological analysis with rabbit anti-lectin IgG and fluorescein-labeled goat anti-rabbit IgG revealed the lectin to be in rat alveolar macrophages and nonparenchymal cells of liver. Thus, the lectin appears to be present in macrophages and is likely involved in receptor-mediated endocytosis. It is distinctly different structurally from the hepatocyte lectin with a similar ligand-binding specificity.

Animals↗
PubMed1979

Qualitative and quantitative aspects of labeling cell surface carbohydrates using lectins as probes.

Lectins are proteins which bind mono- and oligosaccharides with great specificity. Many polysaccharides, glycoproteins and glycolipids which are important constituents of cell walls and surface membranes of prokaryotic and eukaryotic cells, contain sugar moieties with which lectins can interact. As a result lectins have been extensively used for the study of cell surface and membrane structure of their labeling. This paper reviews and evaluates the available methods for the preparation of fluorescent, electron-dense and radioactive lectin derivatives. The procedures involved in the visualization of lectin binding under light and electron microscopy are described. In addition, the methods for quantitative evaluation of the fluorescence on labeled cells and for analysis of the number of cell surface lectin binding sites using radioactively-labeled lectins are outlined. Examples are given for the application of fluorescent lectin derivatives for the detection of specific saccharide-containing molecules on the surfaces of living or fixed microbial cells and various normal and neoplastic cells. The use of lectins to demonstrate the dynamic nature of cell membranes and to detect changes in membrane structure or organization which occur or organization which occur during differentiation, development or after neoplastic transformation is discussed.

Carbohydrates↗
PubMed1986

Purification and characterization of a human lectin specific for penultimate galactose residues.

A novel lectin has been found in human plasma. The lectin was purified by affinity chromatography using an adsorbent in which 2-O-alpha-D-glucopyranosyl-O-beta-D-galactopyranosylhydroxylysine (Glc-Gal-Hyl) was coupled to Sepharose. The molecular weight of the lectin was determined by gradient gel electrophoresis to be approximately 240,000. On polyacrylamide gel electrophoresis in sodium dodecyl sulphate, the subunit had the molecular weight of 29,500. Composition analysis has shown the lectin is a glycoprotein in which 12% of the molecule consists of carbohydrate. Native human, horse, calf, sheep, rabbit, and rat erythrocytes were agglutinated by the lectin in the presence of calcium. Glc-Gal-Hyl, N-acetylated Glc-Gal-Hyl, and stachyose inhibited the hemagglutination, whereas monosaccharides, maltose, cellobiose, lactose, raffinose, galactosylhydroxylysine, and N-acetylated galactosylhydroxylysine were not inhibitory. The lectin is strongly inhibited by the desialylated bovine erythrocyte glycoprotein, which contains galactose beta 1-3galactose beta-sequence at the nonreducing termini of the sugar chains, whereas disialylated orosomucoid did not inhibit the lectin. These results indicate that the lectin recognizes the penultimate galactose residue in a hapten molecule in contrast to usual galactose-binding proteins or galactose-specific lectins, which recognize exposed, terminal galactose residues of sugar chains.

Amino Acids↗