The complement binding-like domains of the murine homing receptor facilitate lectin activity.

The leukocyte homing receptor (HR), the endothelial leukocyte adhesion molecule, and gmp140/platelet activation-dependent granule membrane protein are members of a family of adhesion molecules, termed the lectin cell adhesion molecules (LEC-CAMS) which are unified by a multi-domain structure containing a lectin motif, an epidermal growth factor-like (egf) motif, and variable numbers of a complement binding-like (CB) motif. Previous data have indicated a predominant role for the lectin motif in cell adhesion directed by the LEC-CAMS, although the egf-like domain of the HR may also play a potential role in cell binding. While the role(s) of the CB domains in the LEC-CAMS is currently not understood, they have been hypothesized to act as rigid spacers or stalks for lectin and perhaps, egf domain presentation. In this paper, we analyze the functional characteristics of murine HR-IgG chimeras containing the lectin, lectin plus egf, and lectin plus egf plus CB domains. The Mel 14 mAb, an adhesion blocking antibody which recognizes a conformational determinant in the N-terminus of the HR lectin domain, shows a significantly decreased affinity for a HR construct which lacks the CB motifs, consistent with the possibility that the CB domains are involved with lectin domain structure. In agreement with this conjecture, HR mutants lacking the CB domains show a profound decrease in lectin-specific interaction with the carbohydrate polyphosphomannan ester, suggesting that the changes in Mel 14 affinity for the lectin domain are reflected in lectin functionality. Various assays investigating the interactions between the HR deletion mutants and the peripheral lymph node high endothelium, including cell blocking, immunohistochemical staining, and radioactively labeled ligand binding, all showed that removal of the CB domains results in a lack of HR adhesive function. These results imply that the CB domains of the HR, and, by analogy, the other members of the LEC-CAM family, may play important structural roles involving induction of lectin domain conformation and resultant functionality

High mannose-specific lectin Msl mediates key interactions of the vaginal Lactobacillus plantarum isolate CMPG5300

To characterize the interaction potential of the human vaginal isolate Lactobacillus plantarum CMPG5300, its genome was mined for genes encoding lectin-like proteins. cmpg5300.05_29 was identified as the gene encoding a putative mannose-binding lectin. Phenotypic analysis of a gene knock-out mutant of cmpg5300.05_29 showed that expression of this gene is important for auto-aggregation, adhesion to the vaginal epithelial cells, biofilm formation and binding to mannosylated glycans. Purification of the predicted lectin domain of Cmpg5300.05_29 and characterization of its sugar binding capacity confirmed the specificity of the lectin for high-mannose glycans. Therefore, we renamed Cmpg5300.05_29 as a mannose-specific lectin (Msl). The purified lectin domain of Msl could efficiently bind to HIV-1 glycoprotein gp120 and Candida albicans, and showed an inhibitory activity against biofilm formation of uropathogenic Escherichia coli, Staphylococcus aureus and Salmonella Typhimurium. Thus, using a combination of molecular lectin characterization and functional assays, we could show that lectin-sugar interactions play a key role in host and pathogen interactions of a prototype isolate of the vaginal Lactobacillus microbiota

Purification and properties of a plant Agglutinin

Thesis (M.A.)--Boston UniversityThis study involved work with extracts of the seeds of Bauhinia purpurea alba, in which an N specific lectin was found by Mäkelä (37) and Boyd and McMaster (16). There were two main purposes in mind. First, to see if the lectin could be of practical use as a typing sera; and secondly, to get some insight into its chemical make-up. After working out a purification method and a method for checking the agglutinating power of the lectin, the lectin was tested against several small samples. The writer then tested this lectin against 90 random blood samples. The lectin disagreed with rabbit anti-N in one case -- the lectin typed one MN as an M. There is as yet no known reason for this difference, so it would seem that one could not use the lectin as a typing sera at the present time. The lectin was treated with formaldehyde and lost all its activity as a result. Erythrocytes were treated with an enzyme, ficin, and were then reacted with the lectin. The results showed an increase in the titer of the lectin, and the specificity of the lectin was lost. The freezing of a sample of the lectin for four months resulted in the loss of some of its activity. It remained specific, however. Upon dialyzing the crude extract against saline or water, the substances (Y) that remained behind in the membrane became nonspecific. This showed that there were at least two parts to the extract, one of large molecular weight because it would not pass through the membrane, and the other smaller in size because it would pass through the membrane. The substance that passed through the membrane was called X. By dialyzing the crude extract against distilled water and lyophilizing the distilled water, a substance was found which gave the reaction of a sugar and also seemed to have either or both glucosamine and galactosamine present. Since X showed the presence of reducing sugars and could be shown to inhibit Y against the M site on erythrocytes (see Table 7), it was thought that some sugars should be tested against Y to see if any of them would be able to inhibit Y. The same sugars were also tested against the lectin (Table 5). It was found that none of the sugars had any effect on the lectin; but raffinose, melibiose, and galactose all were found to inhibit Y (Table 8). It has also been shown by other workers (13) that lactose will inhibit Y. The inhibition of the sugars against lectin and Y were run in three ways. The first was the usual inhibition method where the inhibiting substance (sugar, in this case) is placed with the substance to be inhibited (Y) and incubated for one hour. Then the appropriate erythrocyte was added. The second method was to place erythrocytes, Y, and sugar all in the same tube at the same time. The third was to incubate sugar and erythrocytes together and then add Y. The results are given in Table 9. It was found by the writer that Method 2 was the best. This agrees with Krüpe as cited in Mäkelä (37) who states that if an appropriate sugar is added to erythrocytes that are agglutinated by a lectin, the lectin will leave the erythrocytes and pick up the sugar. This is shown by the breaking up of the clumps of erythrocytes. The sugars that would inhibit Y were tried against adsorbed rabbit sera, both anti-M and N. The sugars showed no effect against the rabbit sera. The writer feels that the lectin may not be as specific as the rabbit sera; thus, these inhibiting sugars may only be closely related to the true configuration of the M site

Endogenous mammalian lectin localized extracellularly in lung elastic fibers.

An affinity-purified antibody preparation raised against a beta-galactoside-binding lectin from bovine lung was used to localize a similar lectin in rat lung by immunofluorescence and by electron microscopy after on-grid staining visualized with colloidal gold conjugated second antibody. The endogenous mammalian lectin was found in smooth muscle cells and squamous alveolar epithelial (type I) cells and was concentrated extracellularly in elastic fibers of pulmonary parenchyma and blood vessels. The extracellular localization of this lectin suggests that it, like others, functions by interaction with extracellular glycoconjugates

The distribution of lectins across the phylum Nematoda : a genome-wide search

Nematodes are a very diverse phylum that has adapted to nearly every ecosystem. They have developed specialized lifestyles, dividing the phylum into free-living, animal, and plant parasitic species. Their sheer abundance in numbers and presence in nearly every ecosystem make them the most prevalent animals on earth. In this research nematode-specific profiles were designed to retrieve predicted lectin-like domains from the sequence data of nematode genomes and transcriptomes. Lectins are carbohydrate-binding proteins that play numerous roles inside and outside the cell depending on their sugar specificity and associated protein domains. The sugar-binding properties of the retrieved lectin-like proteins were predicted in silico. Although most research has focused on C-type lectin-like, galectin-like, and calreticulin-like proteins in nematodes, we show that the lectin-like repertoire in nematodes is far more diverse. We focused on C-type lectins, which are abundantly present in all investigated nematode species, but seem to be far more abundant in free-living species. Although C-type lectin-like proteins are omnipresent in nematodes, we have shown that only a small part possesses the residues that are thought to be essential for carbohydrate binding. Curiously, hevein, a typical plant lectin domain not reported in animals before, was found in some nematode species

The serlogical specificity of the lectin from Lens culinaris

Lens culinaris, the common lentil, contains a lectin which has been shown to be specific for a glycoprotein saliva antigen and a glycolipoprotein serum antigen. Both the saliva and serum precipitin reactions with the lectin are directly inhibited with saccharides, especially those related to D-mannose. Electrophoresis of the serum antigen showed that it migrates as three bands, while appearing as a single band in double diffusion precipitin patterns. Quantitative studies of the saliva antigen levels by hemagglutination inhibition titration indicated a polygenic, quantitative mode of inheritance with a minimum heritability of O. 34. Blood group ABH secretor individuals were found to have a significantly lower mean saliva antigen level than nonsecretor individuals. The lectins from Pisum sativum and Canavaliafiensiformis formed precipitin bands of identity with L.culinaris lectin against saliva. C. ensiformis and L. culinaris lectins exhibited precipitin bands of partial identity against serum; and P. sativum and L. culinaris lectins exhibited a pattern of identity against serum. In addition, precipitin patterns of partial identity with the non-H lectin from Lotus tetragonolobus has been demonstrated. Using Ulex europaeus lectin in hemagglutination inhibition experiments with saliva from blood group O secretor individuals, a minimum heritability of approximately 0.40 for H antigen levels was found. A higher frequency of nonsecretor individuals was observed in the Black population compared with the White population

High glucose disrupts oligosaccharide recognition function via competitive inhibition : a potential mechanism for immune dysregulation in diabetes mellitus

Diabetic complications include infection and cardiovascular disease. Within the immune system, host-pathogen and regulatory host-host interactions operate through binding of oligosaccharides by C-type lectin. A number of C-type lectins recognise oligosaccharides rich in mannose and fucose – sugars with similar structures to glucose. This raises the possibility that high glucose conditions in diabetes affect protein-oligosaccharide interactions via competitive inhibition. Mannose binding lectin, soluble DC-SIGN & DC-SIGNR, and surfactant protein D, were tested for carbohydrate binding in the presence of glucose concentrations typical of diabetes, via surface plasmon resonance and affinity chromatography. Complement activation assays were performed in high glucose. DC-SIGN and DC-SIGNR expression in adipose tissues was examined via immunohistochemistry. High glucose inhibited C-type lectin binding to high-mannose glycoprotein and binding of DC-SIGN to fucosylated ligand (blood group B) was abrogated in high glucose. Complement activation via the lectin pathway was inhibited in high glucose and also in high trehalose - a nonreducing sugar with glucoside stereochemistry. DC-SIGN staining was seen on cells with DC morphology within omental and subcutaneous adipose tissues. We conclude that high glucose disrupts C-type lectin function, potentially illuminating new perspectives on susceptibility to infectious and inflammatory disease in diabetes. Mechanisms involve competitive inhibition of carbohydrate-binding within sets of defined proteins, in contrast to broadly indiscriminate, irreversible glycation of proteins

Lectin-like bacteriocins from pseudomonas spp. utilise D-rhamnose containing lipopolysaccharide as a cellular receptor

Lectin-like bacteriocins consist of tandem monocot mannose-binding domains and display a genus-specific killing activity. Here we show that pyocin L1, a novel member of this family from Pseudomonas aeruginosa, targets susceptible strains of this species through recognition of the common polysaccharide antigen (CPA) of P. aeruginosa lipopolysaccharide that is predominantly a homopolymer of d-rhamnose. Structural and biophysical analyses show that recognition of CPA occurs through the C-terminal carbohydrate-binding domain of pyocin L1 and that this interaction is a prerequisite for bactericidal activity. Further to this, we show that the previously described lectin-like bacteriocin putidacin L1 shows a similar carbohydrate-binding specificity, indicating that oligosaccharides containing d-rhamnose and not d-mannose, as was previously thought, are the physiologically relevant ligands for this group of bacteriocins. The widespread inclusion of d-rhamnose in the lipopolysaccharide of members of the genus Pseudomonas explains the unusual genus-specific activity of the lectin-like bacteriocins

Beyond the fuzzy lock-and-key: spontaneous symmetry shifts and glycan/lectin logic gates

Changes in the molecular topology of glycan/lectin interaction may explain observed reaction punctuation driven by experimental gradients in reactant concentration. Adoption of a 'biological renormalization' perspective from statistical physics for the analysis of such phase transitions suggests, in marked contrast to conventional physical systems, a broad spectrum of possible universality class behaviors. This spectrum may, in typical perverse biological manner, be of central scientific interest. Generalization, via formalism abducted from coevolutionary theory, suggests that glycan/lectin molecular switches instantiate logic gates that may be as sophisticated as those characterizing basic neural process, if on a different scale