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PubMed1993

Immobilization of Fc epsilon receptors by wheat germ agglutinin. Receptor dynamics in IgE-mediated signal transduction.

Transmembrane signaling initiated by the receptor with high affinity for Fc stem of IgE(Fc epsilon RI) requires the diffusion-dependent cross-linkage and persistent aggregation of the Fc epsilon RI. Disruption or prevention of receptor cross-links at any time during the secretory response quickly terminates secretion. We found that in the rat basophilic leukemia mast cell line, addition of wheat germ agglutinin, a lectin that binds to the Fc epsilon RI alpha subunit, caused a precipitous decline in the lateral diffusional and electrokinetic mobilities of the Fc epsilon RI. Both the unoccupied Fc epsilon RI and IgE-Fc epsilon RI complexes became immobilized, as determined from in situ electromigration and postelectric field relaxation. Immobilization of the Fc epsilon RI by wheat germ agglutinin was accompanied by a ligand-reversible association of 125I-IgE-Fc epsilon RI complexes with the Triton X-100-insoluble cytoskeletal fraction. Wheat germ agglutinin rapidly inhibited Fc epsilon RI-mediated signal transduction and secretion, whether cross-linkage was initiated by multivalent antigen, covalent IgE oligomers, anti-IgE, or anti-Fc epsilon RI antibody. Inhibition of signaling and secretion occurred on simultaneous addition of wheat germ agglutinin and antigen, and also when wheat germ agglutinin was added at increasing times after induction of Fc epsilon RI cross-linkage. Wheat germ agglutinin neither reduced the affinity of anti-DNP IgE for haptenic DNP-lysine nor reversed the binding of IgE to the Fc epsilon RI. Although wheat germ agglutinin caused internalization of the Fc epsilon RI, the onset of inhibition preceded and its extent exceeded that of internalization. Wheat germ agglutinin did not interfere with the secretory apparatus, as indicated by its lack of inhibition of secretion elicited by calcium ionophores. These findings suggest that inhibition of signal transduction is secondary to an initial event linked to immobilization of the Fc epsilon RI. Implications of these results are discussed with respect to the dynamics of Fc epsilon RI aggregation on rat basophilic leukemia cells.

Animals↗

PubMed1991

Transmembrane cytoskeletal modulation in preterminal growing axons. II. Limax flavus agglutinin-induced receptor redistribution, capping and internalization in varicosities of growing axons.

Growing retinal ganglion cell axons of the goldfish exhibit varicosities of varying sizes and smaller non-protruding phase-dense inclusions that are mobile and mediate rapid bulk redistribution of axoplasm. In fixed axons, Limax flavus agglutinin, a lectin specific for sialic acid which has been shown to inhibit organelle transport in these axons, preferentially labels surface membrane associated with varicosities and inclusions in preterminal axons. In viable axons, Limax flavus agglutinin causes: (1) agglutination of closely apposed axons, (2) redistribution of lectin-binding sites on varicosities to surfaces of interaxonal contact with other varicosities, forming 'fused' multivaricosity complexes, and (3) formation of vacuoles in many single varicosities and some multivaricosity complexes. Vacuoles contain Limax flavus agglutinin binding sites distributed circumferentially. On the basis of immunocytochemistry, actin, myosin, calmodulin and alpha-spectrin are co-localized with redistributed Limax flavus agglutinin binding sites. The agglutination, redistribution of lectin binding sites and changes in the cytoskeleton can be reversed by treatment with sialic acid. The lectin-induced vacuole formation and internalization of Limax flavus agglutinin receptors can also be blocked either by sodium azide in a glucose-free medium, or by pretreatment with cytochalasin D and indicate an energy and a cytoskeletal dependence. The Limax flavus agglutinin-induced structural rearrangements are not altered after limited digestion with pronase. Western blots after ultramicroelectrophoresis of retinal ganglion cell axons subjected to limited digestion reveal Limax flavus agglutinin labelling of bands with apparent Mr of 64 and 70 KDa. In undigested axons, some 70 KDa protein remains unextracted with Triton X-100 lysis of axonal fields, and more remains unextracted when axonal fields are pretreated with Limax flavus agglutinin before Triton lysis, suggesting increased association with the cytoskeleton in response to lectin binding. The results indicate that cross-linking of one or more sialoglycoconjugates on the surface of varicosities of preterminal growing retinal ganglion cell axons causes a constellation of transmembrane-mediated cytoskeletal and membrane changes that are akin to those described for capping in motile cells.

Actins↗

PubMed1984

Chlamydomonas agglutinin conjugated to agarose beads as an in vitro probe of adhesion.

Flagellar sexual agglutinins are responsible for the primary recognition and adhesion events of mating in Chlamydomonas reinhardi which culminate in zygotic union of plus and minus gametes. Recent studies in this laboratory have shown the plus agglutinin to be an extremely large (greater than 10(6) D) and asymmetric glycoprotein containing a high proportion of hydroxyproline and serine residues [14, 27, 28]. This paper reports an improved method for in vitro investigations of the adhesive nature of this molecule. Purified agglutinin is covalently attached to an insoluble (Affi-gel 15 agarose bead) support and shown to retain potent agglutination activity when presented to living minus gametes, which rapidly and extensively adhere to the coated bead surface by their flagella. The specificity of the response is documented by the lack of interaction of plus gametes with the immobilized plus agglutinin (IA+). Using this simple yet sensitive bioassay, we have subjected IA+ beads to various enzymatic, chemical and physical treatments and assessed the effects on agglutinin activity. These studies reveal that Chlamydomonas plus agglutinin is sensitive to thermolysin or trypsin digestion, alkaline borohydride reduction, periodate oxidation, thiol reduction and heating at 65 degrees C, but unaffected by treatment with chymotrypsin, endo- or exoglycosidases, or incubation with isolated minus agglutinin. The implications of these results for agglutinin structure and possible functional interactions in initial recognition/adhesion events are discussed.

Adhesiveness↗

PubMed1989

The plant lectin soybean agglutinin binds to the soma, axon and central terminals of a subpopulation of small-diameter primary sensory neurons in the rat and cat.

The distribution of binding of the plant lectin soybean agglutinin to primary sensory neurons has been investigated in the rat and cat. Soybean agglutinin was found to bind to approximately 30% of neurons in the fourth, lumbar dorsal root ganglion of the rat and approximately 50% of neurons in the first, sacral dorsal root ganglion in the cat. Morphometric analysis of these dorsal root ganglia revealed that in both species those neurons which bind soybean agglutinin appear to be a subpopulation of the small-diameter cells. Electron microscopic analysis of the lumbar dorsal roots revealed that soybean agglutinin binds to the plasma membrane of a subpopulation of unmyelinated axons in both species. Myelinated axons did not bind the lectin. In the dorsal horn of the spinal cord, soybean agglutinin bound to components of Lissauer's tract and the superficial laminae (laminae I and II) of the dorsal horn in both species. In the dorsal horn lectin binding was limited to lamina I and the outer portion of lamina II (sublamina IIo) in the cat, while in the rat lamina I and the entire dorsoventral extent of lamina II (sublaminae IIo and IIi) were labelled. Thirty days following dorsal rhizotomy, soybean agglutinin binding in the superficial dorsal horn, ipsilateral to the rhizotomy, was abolished in both species. These results show the plant lectin soybean agglutinin to be a histochemical marker for the soma and central terminals of a subpopulation of small-diameter sensory neurons in both the rat and cat. It is suggested that soybean agglutinin binding may be used in conjunction with immunohistochemistry to allow identification of putative neurotransmitters within, or in synaptic profiles associated with, the central terminals of small-diameter primary sensory neurons.

Animals↗

PubMed1997

The complex-type oligosaccharide binding lectin Datura stramonium agglutinin detects type II A muscle fibres in the branchial biceps from man and cat.

Complex-type oligosaccharides were detected in the sarcoplasm of muscle fibres from cat and human biceps using lectins and anticarbohydrate antibodies. The lectin Datura stramonium agglutinin strongly stained type II A fibres as identified by myosin ATPase activity after alkaline and acid preincubation. In contrast, all muscle fibres showed a moderate coarse granular staining after incubation with Tetracarpidum conophorum agglutinin and Telfairia occidentalis agglutinin which recognize tri-antennary complex glycans poorly bound by D. stramonium agglutinin. Strong sarcoplasmic staining in all muscle fibres was obtained after incubation with an antibody against branched N-acetyllactosamine structure while an antibody against binary 2 --> 3 sialyllactosamine glycans failed to detect the muscle fibres. Treatment of the muscle sections with sialidase prior to incubation with D. stramonium agglutinin did not influence the lectin staining pattern. Staining of blots from electrophoretically separated muscle proteins obtained by homogenization, solubilization and centrifugation of small muscle pieces showed D. stramonium agglutinin binding to a number of bands ranging from 200 kDa to 30 kDa. No D. stramonium agglutinin positive bands were observed in blots from separated mitochondrial proteins while blots from sarcoplasmic reticulum separated by electrophoresis stained many bands in the range from 200 kDa to 30 kDa. It may be concluded that all muscle fibres in human and cat biceps hold intracellular non-sialylated complex-type oligosaccharides and further, that a specific tri-antennary complex-type glycoform is strongly expressed in type II A fibres as recognized by D. stramonium agglutinin. These results indicate a different glycosylation of certain myofibrillar-associated proteins in muscle fibre types.

Animals↗

PubMed2006

Effects of soybean agglutinin on body composition and organ weights in rats.

An experiment was conducted to assess the effect of soybean agglutinin dosage level on growth, body composition, plasma lipids, glucose, urea nitrogen content and aminotransferase activities in rats. Male and female rats (n=60) weaned at 19 d were given a dose of 0, 3.5, 7.0, 10.5, or 14.0 mg soybean agglutinin by gastric infusion once daily for 10 days. With increasing doses of soybean agglutinin, body weight, lipid content of carcass, spleen and kidneys relative dry weights decreased, while small intestine and pancreatic weight, the contents of urea nitrogen and triglyceride, and the activities of aspartate aminotransferase linearly increased in plasma. Though soybean agglutinin decreased plasma insulin content, changes in plasma glucose content due to soybean agglutinin were not detected. It is suggested that dietary soybean agglutinin may affect the secretion of other hormones besides insulin, which modulate blood glucose reserves. In conclusion, consumption of soybean agglutinin resulted in a depletion of lipid and an overgrowth of small intestine and pancreas in rats. Meanwhile, poor growth of spleen and kidneys was observed in the soybean agglutinin-fed rats.

Animal Feed↗

PubMed1992

Recognition of collagen by fibroblasts through cell surface glycoproteins reactive with Phaseolus vulgaris agglutinin.

The role of glycochains of cell surface glycoproteins in the cell to collagen interaction was examined by studying the effect of lectins on the fibroblast-mediated collagen gel contraction. Lectins of Phaseolus vulgaris agglutinin (PHA), concanavalin A (ConA), lentil seed agglutinin (LCA), pea agglutinin (PSA), Ricinus communis agglutinin-60 (RCA), and wheat germ agglutinin (WGA) dose-dependently inhibited gel contraction, while lectins of mushroom agglutinin (ABA), peanut agglutinin (PNA), pokeweed mitogen (PWM), and soybean agglutinin (SBA) did not. Of these lectins, PHA seemed to be worthy of further analysis, because PHA, but not other lectins, inhibited spreading of fibroblasts on collagen fibrils but not on plastic or gelatin, suggesting that cell-surface glycoproteins responsive to the lectin are involved in the specific binding of fibroblasts to native collagen fibrils. The inhibitory effect of PHA-E4, an isolectin of PHA, was more intense than that of PHA-L4, another isolectin of PHA. The collagen gel contraction was also inhibited by tunicamycin and monensin in a concentration-dependent and reversible manner. These results strongly suggest that PHA-E4-reactive glycoproteins of the fibroblast surface play an important role in cell to collagen binding during the gel contraction. Five membrane proteins including beta 1 subunits of the integrin family were obtained by affinity chromatography with PHA-E4.

Cell Movement↗

PubMed1986

Interaction of sialoglycoproteins with wheat germ agglutinin-sepharose of varying ratio of lectin to Sepharose. Use for the purification of mucin glycoproteins from membrane extracts.

The influence of varying the amount of wheat germ agglutinin immobilized on Sepharose beads on the binding of glycoproteins to these beads was investigated. A series of wheat germ agglutinin-Sepharose gels containing between 0.10 and 10.0 mg of lectin/ml of gel was prepared, and the actual lectin content was established by acid hydrolysis of the gel followed by analysis of glycine, a major amino acid in wheat germ agglutinin. Affinity chromatography of labeled glycoproteins indicated that glycophorin bound to all the wheat germ agglutinin-Sepharose preparations. Fetuin, ovomucoid, and alpha 1-acid glycoprotein bound not at all or very poorly to gels with a low content of wheat germ agglutinin (less than 0.95 mg/ml). The specific binding of these glycoproteins increased with increasing lectin content on the gels, and on gels of high content (greater than 3 mg/ml) the binding was virtually quantitative. On chromatographing a mixture of glycophorin, alpha 1-acid glycoprotein, fetuin, and ovomucoid on wheat germ agglutinin-Sepharose, containing 0.08 mg of lectin/ml of gel, glycophorin was selectively retained on the gel. It was possible to purify glycophorin from an extract of human erythrocyte membranes in one step by chromatography on the above gel. By using the series of gels, it was demonstrated that Morris hepatoma 7777 membranes contained at least 4-fold more sialoglycoproteins which bound to low density wheat germ agglutinin-Sepharose compared to rat liver membranes. These hepatoma sialoglycoproteins were isolated, purified, and partially characterized as having a high proportion of O-linked sialyloligosaccharides. Our studies illustrate the use of low density wheat germ agglutinin-Sepharose gels both for the detection and for easy isolation of mucin-type glycoproteins from crude extracts of cells or membranes.

Animals↗

PubMed1994

Transganglionic labelling of primary sensory afferents in the rat lumbar spinal cord: comparison between wheatgerm agglutinin and the I-B4 isolectin from Bandeiraea simplicifolia.

We recently reported that the I-B4 isolectin from Bandeiraea simplicifolia could be used as a transganglionic neuronal tracer which appears to be selective for unmyelinated cutaneous afferents (C fibres) and their terminals in the superficial dorsal horn. As terminals in the superficial dorsal horn are also labelled by wheatgerm agglutinin, we sought to compare these two neuronal tracers. Three days after the injection of 1% wheatgerm agglutinin-HRP or 1% BSI-B4-HRP into the sciatic nerve of adult rats the lumbar spinal cord was processed for HRP reactivity. The majority of labelled structures was found in the superficial dorsal horn, with fewer labelled structures seen in the overlying white matter (including Lissauer's tract). In wheatgerm agglutinin-HRP experiments most labelled structures were synaptic terminals (63%) and unmyelinated axons (32%). About 3% of wheatgerm agglutinin-HRP-labelled structures were fine myelinated fibres (which were found only in lamina I and outer lamina II) and about 2% of label was located in neuronal somata. In contrast, label from BSI-B4-HRP experiments was found only in synaptic terminals (37%) and unmyelinated axons (63%). Previous studies have shown that small diameter dorsal root ganglion neurons and their terminals in the superficial dorsal horn express a range of structurally related carbohydrates that contain binding sites for BSI-B4 or wheatgerm agglutinin or both. Comparison of the labelling patterns produced by the two transganglionic tracers in the present study suggests that unmyelinated sciatic afferents express wheatgerm agglutinin and BSI-B4 binding sites, but some thin myelinated afferents, and a distinct form of synaptic terminal in lamina I/II outer, express the wheatgerm agglutinin binding site and not the BSI-B4 binding site.

Afferent Pathways↗

PubMed1999

Anti-T antibodies and peanut-agglutinin-binding glycoproteins in sera of patients with gastric cancer.

Agglutinating antibodies to neuraminidase-treated red blood cells (anti-T agglutinins) are known to be reduced in patients with gastric cancer. The antigenic determinant of anti-T agglutinin is known to have a disaccharide structure [Gal(beta1-3)GalNAc], the same specificity as peanut agglutinin (PNA). We examined sera of 27 patients with gastric cancer and 30 controls for anti-T agglutinins, anti-T antibodies and PNA-binding glycoproteins. Anti-T agglutinins were titrated by a microtiter hemagglutination method. Levels of anti-T antibodies were determined by enzyme immunoassay using synthetic glycoconjugate [Gal(beta1-3)GalNAc O-alpha-linked to human serum albumin] as an antigen. Levels of PNA-binding glycoproteins in sera were measured by sandwich enzyme-linked lectin assay using wheat germ agglutinin and peroxidase-conjugated PNA. Titers of anti-T agglutinins were significantly lower in patients with gastric cancer than in controls (P = 0.041). Levels of anti-T antibodies were not significantly different in patients with gastric cancer and controls; however, decreased levels of anti-T antibodies were more frequent in patients with gastric cancer than in controls (P = 0. 001). Levels of PNA-binding glycoproteins were significantly higher in sera of patients with gastric cancer than in controls (P = 0.001). The levels of anti-T antibodies inversely correlated with the levels of PNA-binding glycoproteins in sera of patients with gastric cancer (r = -0.44, P = 0.021). These results suggest that the decrease in anti-T antibodies in sera of patients with gastric cancer might be due to immune complex formation between circulating PNA-binding glycoproteins and anti-T antibodies.

Adult↗

PubMed2000

Biochemical characterization of sialoprotein "anti-agglutinin" purified from boar epididymal and seminal plasma.

Sialoprotein "anti-agglutinin," previously shown to inhibit sperm head-to-head agglutination, is found in both boar epididymal and seminal plasma. The present report characterizes anti-agglutinin by mass spectrometry, by N-terminal amino acid sequence analysis, and by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE) and Western blotting techniques to assess phosphate content of the molecule. Anti-agglutinin had the SDS-PAGE mobility of approximately 25 kDa. By electrospray ionization-mass spectrometry, however, mass spectra of anti-agglutinin were characterized by two major peaks (19,379-19,382 Da and 19,395-19,397 Da) and several minor peaks. Mass spectrometry of tryptic peptide fragments of deglycosylated anti-agglutinin and amino acid sequence analysis revealed that the protein has a unique peptide-mass fingerprinting of fragments (12,668 Da, 5,209 Da, 1,226 Da, and 1,168 Da) and a novel N-terminal amino acid sequence (KTDDY AISGA KEEEF YDYME ELYAV), respectively. Additionally Western blot techniques, using commercially available monoclonal antibodies, were used to detect presence of phosphothreonine and phosphoserine substituents, but two different monoclonal antibodies did not detect phosphotyrosine. Moreover, treatment with two different alkaline phosphotases converted the molecule, as assessed by SDS-PAGE and detection by silver stain, from the parent form of about 25 kDa to forms of approximately 19 kDa (similar to that assigned by mass spectrometry) and/or 15 kDa. Original antiserum generated toward, and reacting with native anti-agglutinin, reacted only with 19 kDa form. These results are consistent with the conclusion that the native anti-agglutinin may be a novel protein that is phosphorylated at serine and/or threonine residues.

Agglutinins↗

PubMed1994

Mating type-specific cell-cell recognition of Saccharomyces cerevisiae: cell wall attachment and active sites of a- and alpha-agglutinin.

Mating type-specific agglutination of Saccharomyces cerevisiae a and alpha cells depends on the heterophilic interaction of two cell surface glycoproteins, the gene products of AG alpha 1 and AGA2. Evidence is presented with immunogold labelling that the alpha-agglutinin is part of the outer fimbrial cell wall coat. The a-agglutinin is bound via two S-S bridges (Cys7 and Cys50) to a cell wall component, most probably the gene product of AGA1. His273 of alpha-agglutinin has previously been shown to be essential for a- and alpha-agglutinin interaction and a model based on two opposing ion-pairs had been proposed. By site-directed mutagenesis this possibility has now been excluded. With the help of various peptides, either chemically synthesized, obtained by proteolysis of intact glycosylated a-agglutinin or prepared from a fusion protein expressed in Escherichia coli, the biologically active region of a-agglutinin was located at the C-terminus of the molecule. A peptide consisting of the C-terminal 10 amino acids (GSPIN-TQYVF) was active in nanomolar concentrations. Saccharide moieties, therefore, are not essential for the mating type-specific cell-cell interaction; glycosylated peptides are, however, four to five times more active than non-glycosylated ones. Comparisons of the recognition sequences of the S. cerevisiae agglutinins with that of the Dictyostelium contact site A glycoprotein (gp80), as well as with those of the various families of cell adhesion molecules of higher eucaryotes, have been made and are discussed.

Agglutinins↗

PubMed2003

Purification of saliva agglutinin of Streptococcus intermedius and its association with bacterial aggregation and adherence.

Streptococcus intermedius strain 1208-1 cells were aggregated in the presence of saliva. The saliva agglutinin was purified by centrifugation, filtration, and gel filtration. SDS-PAGE analyses indicated that the purified agglutinin consisted of two high-molecular-mass proteins. Aggregation was dependent on calcium over pH 5.5, with 1 mM being the most effective concentration. Boiling inactivated purified agglutinin. S. intermedius strain 3 and Streptococcus mutans strain 1 were aggregated in the purified agglutinin. After adsorption with strain 1208-1 cells, the saliva sample did not exhibit any aggregation activity, and the agglutinin bands were no longer visible by SDS-PAGE. Adherence analyses demonstrated that the purified agglutinin immobilized on the surfaces of polystyrene wells, actinomyces cells, and apatite beads accounted for the binding of streptococcus cells. Agglutinin also effectively inhibited adherence to apatite beads coated with native saliva.

Actinomyces↗

PubMed1983

N-[acetyl-3H] wheat germ agglutinin: anatomical and biochemical studies of a sensitive bidirectionally transported axonal tracer.

Both retrograde and anterograde autoradiographic axonal labeling were associated with the appropriate pathways after injections of N-[acetyl-3H] wheat germ agglutinin within numerous structures in the central nervous system of mice. Sections processed for light microscopic autoradiography following the placement of injections mainly within the neocortex, neostriatum, or cerebellum have revealed patterns of bidirectional axonal labeling in various thalamic, monoaminergic, deep cerebellar and precerebellar nuclei that were similar to that seen after similarly placed horseradish peroxidase injections. Small injections of N-[acetyl-3H] wheat germ agglutinin, by way of an extremely limited extracellular spread of the tracer, yield large amounts of autoradiographic retrograde and anterograde axonal labeling. High specific activities, in addition to the sensitivity displayed by this tracer, allow the use of relatively short autoradiographic exposure times that still lead to an extensive signal over labeled neurons. Biochemical analysis of this radiolabeled derivative of wheat germ agglutinin was carried out using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Results from Coomassie blue staining and fluorography of the gels have revealed that affinity-purified N-[acetyl-3H] wheat germ agglutinin migrates predominantly as a monomer (molecular weight 18,000 +/- 1000 daltons) as well as a dimer (35,100 +/- 1000 daltons) when compared with native wheat germ agglutinin and molecular weight standards. Gels run on the native preparation, prior to derivatization, radiolabeling and affinity purification, in addition to containing species that co-migrate with the monomeric and dimeric forms of the tagged lectin, also reveal several other bands with molecular weights above and below those values for the monomer and dimer. When scintillation counting was performed on fresh gels of N-[acetyl-3H] wheat germ agglutinin, the predominant species was again found to be the monomeric and dimeric forms; however, small peaks were also present around 12,000 daltons as well as above 43,000 daltons. Coomassie blue staining and fluorography never revealed banding below 18,000 daltons, though some trace of labeling was present, but not in the form of discrete bands, above 36,000 daltons. The presence, albeit small in comparison to the monomer and dimer, of these other higher and lower molecular weight species might in part result from fragments produced by proteolytic digestion of intact subunits of N-[acetyl-3H] wheat germ agglutinin that may or may not possess intact binding sites.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1990

Comparison of Streptococcus mutans and Streptococcus sanguis receptors for human salivary agglutinin.

Oral streptococci vary in their susceptibility to salivary agglutinin-mediated aggregation. To understand the molecular basis of this specificity, the structure and function of receptors for agglutinin from Streptococcus mutans KPSK2 (MSL-1) and Streptococcus sanguis M5 (SSP-5) were compared. Immunological screening of an S. mutans KPSK2 genomic DNA library yielded two identical clones expressing a streptococcal protein that co-migrated with a 220 kDa peptide in SDS extracts from this organism. This protein inhibited agglutinin-mediated aggregation of S. mutans KPSK2 in a dose-dependent manner. The MSL-1 gene is homologous to the S. mutans SpaP and pac genes although single base substitutions alter several amino acids. MSL-1 is also similar to the agglutinin receptor (SSP-5) cloned from S. sanguis M5. All three proteins, MSL-1, P1, and SSP-5 share at least one epitope since monoclonal and polyclonal anti-SSP-5 antibodies react with both MSL-1 and P1. However, other monoclonal antibodies are specific for SSP-5 and appear to react with a peptide domain exhibiting little homology to MSL-1 or P1. Sugar inhibition studies showed that agglutinin-mediated aggregation of S. mutans KPSK2 was most potently inhibited by fucose and lactose. Sialic acid, a potent inhibitor of S. sanguis aggregation, had no effect on the interaction of agglutinin with S. mutans KPSK2. These results suggest that while the MSL-1 and SSP-5 proteins are genetically and immunologically related, their specificity for binding sites on agglutinin differs.

Adhesins, Bacterial↗

PubMed1999

Purification and characterization of a natural agglutinin from the serum of the hermit crab Diogenes affinis.

A natural agglutinin from the serum of the hermit crab Diogenes affinis was purified to homogeneity by a single-step affinity chromatography using N-acetylglucosamine-coupled Sepharose 6B. The purified serum agglutinin (PSA) showed a strong affinity for rat RBC, and its hemagglutinating (HA) activity was specifically dependent on Ca2+ and reversibly sensitive to EDTA. PSA in active form has a molecular mass estimate of 185 kDa and is composed of four non-identical subunits (51, 49, 42 and 39 kDa) cross-linked by interchain disulfide bonds. The homogeneity of PSA was corroborated by immunodiffusion and immunoelectrophoretic analyses using rabbit antiserum raised against the agglutinin. The antibodies in this antiserum appear to be specific for RBC-binding sites of the agglutinin molecules as revealed by the ability of the antiserum to neutralize HA activities of both whole serum and PSA of D. affinis. In HA-inhibition assays performed with several carbohydrates and glycoproteins, PSA showed a distinct and unique specificity for acetyl group in carbohydrates independently of the presence of this group on C-2 or C-5 and its stereochemical arrangement in the axial or equatorial orientation. Besides, this agglutinin appears to recognize the terminal N- and O- acetyl groups in the oligosaccharide chain of glycoconjugates. The HA activity of D. affinis agglutinin was also susceptible to inhibition by lipopolysaccharides from diverse gram-negative bacteria, which might indicate a significant in vivo role of this humoral agglutinin in the host immune response against bacterial infections.

Agglutinins↗

PubMed1981

Receptor-mediated gonadotropin action in the ovary. Inhibitory actions of concanavalin A and wheat-germ agglutinin on gonadotropin-stimulated cyclic AMP and progesterone responses in ovarian cells.

Pretreatment of ovarian cells with concanavalin A and wheat-germ agglutinin blocked the gonadotropin-induced cyclic AMP and progesterone responses and this effect was time- and concentration-dependent. Basal production of either cyclic AMP or progesterone, however, was not affected by treatment of cells with lectin. The effect of concanavalin A on gonadotropin-mediated cyclic AMP and progesterone responses was blocked by alpha-methyl D-mannoside and alpha-methyl d-glucoside. Similarly the inhibitory effect of wheat-germ agglutinin was reversed by N-acetyl-D-glucosamine. Pretreatment of ovarian cells with concanavalin A or wheat-germ agglutinin had no effect on protein synthesis in the ovary as monitored by [3H]proline incorporation studies. Concanavalin A and wheat-germ agglutinin did not affect steroid production in response to dibutyryl cyclic AMP and 8-bromo cyclic AMP, indicating that the inhibitory action of lectin was occurring at a step before cyclic AMP formation. Lectins specific for L-fucose, D-galactose and N-acetyl-D-galactosamine, gorse seed agglutinin, peanut agglutinin and Dolichos biflorus agglutinin respectively, did not interfere with gonadotropin-induced cyclic AMP and progesterone responses. The present studies suggest that gonadotropin receptors may be glycoprotein in nature or closely associated with glycoprotein structures with the carbohydrate chain containing N-acetyl-D-glucosamine, mannose and possibly N-acetylneuraminic acid.

Animals↗

PubMed1987

Studies on chemical modification of cold agglutinin from the snail Achatina fulica.

The cold agglutinin isolated from the albumin gland of the snail Achatina fulica was modified with various chemical reagents in order to detect the amino acids and/or carbohydrate residues present in its carbohydrate-binding sites. Treatment with reagents considered specific for modification of lysine, arginine and tryptophan residues of the cold agglutinin did not affect the carbohydrate-binding activity of the agglutinin. Modification of tyrosine residues showed some change. However, modification with carbodiimide followed by alpha-aminobutyric acid methyl ester causes almost complete loss of its binding activity, indicating the involvement of aspartic acid and glutamic acid in its carbohydrate-binding activity. The carbohydrate residues of the cold agglutinin were removed by beta-elimination reaction, indicating that the sugars are O-glycosidically linked to protein part of the molecule. Removal of galactose residues from the cold agglutinin by the action of beta-galactosidase indicated that the galactose molecules are beta-linked. These carbohydrate-modified glycoproteins showed a marked change in agglutination property, i.e. they agglutinated rabbit erythrocytes at both 10 degrees C and 25 degrees C, indicating that the galactose residues of the glycoprotein play an important role in the cold-agglutination property of the glycoprotein. The c.d. data showed the presence of an almost identical type of random-coil conformation in the native cold agglutinin at 10 degrees C and in the carbohydrate-modified glycoprotein at 10 degrees C and 25 degrees C. This particular random-coil conformation is essential for carbohydrate-binding property of the agglutinin.

Agglutinins↗