Lectins: carbohydrate-specific reagents and biological recognition molecules.
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Animals↗
Biomedical subjects
Nathan Sharon
Publications and source records attributed to Nathan Sharon.
Carbohydrates as future anti-adhesion drugs for infectious diseases.
Adhesion of pathogenic organisms to host tissues is the prerequisite for the initiation of the majority of infectious diseases. In many systems, it is mediated by lectins present on the surface of the infectious organism that bind to complementary carbohydrates on the surface of the host tissues. Lectin-deficient mutants often lack the ability to initiate infection. The bacterial lectins are typically in the form of elongated submicroscopic multi-subunit protein appendages, known as fimbriae (or pili). The best characterized of these are the mannose-specific type 1 fimbriae, the galabiose-specific P fimbriae and the N-acetylglucosamine-specific fimbriae of Escherichia coli. Soluble carbohydrates recognized by the bacterial surface lectins block the adhesion of the bacteria to animal cells in vitro. Aromatic alpha-mannosides are potent inhibitors of type 1 fimbriated E. coli, being up to 1000 times more active than MealphaMan, with affinities in the nanomolar range. This is due to the presence of a hydrophobic region next to the monosaccharide-binding site of the fimbriae, as recently demonstrated by X-ray studies. Polyvalent saccharides (e.g., neoglycoproteins or dendrimers) are also powerful inhibitors of bacterial adhesion in vitro. Very significantly, lectin-inhibitory saccharides have been shown to protect mice, rabbits, calves and monkeys against experimental infection by lectin-carrying bacteria. Since anti-adhesive agents do not act by killing or arresting the growth of the pathogens, it is very likely that strains resistant to such agents will emerge at a markedly lower rate than of strains that are resistant to antibiotics. Suitable sugars also inhibit the binding to cells of carbohydrate-specific toxins, among them those of Shigella dysenteriae Type 1, and of the homologous Verotoxins of E. coli, specific for galabiose. Appropriately designed polyvalent ligands are up to six orders of magnitude stronger inhibitors of toxin binding in vitro than the monovalent ones, and they protect mice against the Shigella toxin. The above data provide clear proof for the feasibility of anti-adhesion therapy of infectious diseases, although this has not yet been successful in humans. All in all, however, there is little doubt that inhibitors of microbial lectins will in the near future join the arsenal of drugs for therapy of infectious diseases.
Animals↗
Meet the multifunctional and sexy glycoforms of glycodelin.
Glycodelin, a human-secreted glycoprotein that appears in a small number of glycoforms, exhibits diverse biological activities, such as in contraception and immunosuppression. Moreover, different tissue-specific glycoforms appear to mediate diverse functions. Quite unusually, the glycodelin N-linked glycans differ between the male and female glycoforms. The fact that these glycans are fundamental for exerting the physiological activities of the different glycoforms, makes them an interesting target for glycobiology research. This review will focus on the involvement of the glycans in glycodelin activity and compare between the several glycoforms.
Animals↗
The distinct binding specificities exhibited by enterobacterial type 1 fimbriae are determined by their fimbrial shafts.
Type 1 fimbriae of enterobacteria are heteropolymeric organelles of adhesion composed of FimH, a mannose-binding lectin, and a shaft composed primarily of FimA. We compared the binding activities of recombinant clones expressing type 1 fimbriae from Escherichia coli, Klebsiella pneumoniae, and Salmonella typhimurium for gut and uroepithelial cells and for various soluble mannosylated proteins. Each fimbria was characterized by its capacity to bind particular epithelial cells and to aggregate mannoproteins. However, when each respective FimH subunit was cloned and expressed in the absence of its shaft as a fusion protein with MalE, each FimH bound a wide range of mannose-containing compounds. In addition, we found that expression of FimH on a heterologous fimbrial shaft, e.g. K. pneumoniae FimH on the E. coli fimbrial shaft or vice versa, altered the binding specificity of FimH such that it closely resembled that of the native heterologous type 1 fimbriae. Furthermore, attachment to and invasion of bladder epithelial cells, which were mediated much better by native E. coli type 1 fimbriae compared with native K. pneumoniae type 1 fimbriae, were found to be dependent on the background of the fimbrial shaft (E. coli versus K. pneumoniae) rather than the background of the FimH expressed. Thus, the distinct binding specificities of different enterobacterial type 1 fimbriae cannot be ascribed solely to the primary structure of their respective FimH subunits, but are also modulated by the fimbrial shaft on which each FimH subunit is presented, possibly through conformational constraints imposed on FimH by the fimbrial shaft. The capacity of type 1 fimbrial shafts to modulate the tissue tropism of different enterobacterial species represents a novel function for these highly organized structures.
Animals↗
History of lectins: from hemagglutinins to biological recognition molecules.
The occurrence in nature of erythrocyte-agglutinating proteins has been known since the turn of the 19th century. By the 1960s it became apparent that such proteins also agglutinate other types of cells, and that many of them are sugar-specific. These cell-agglutinating and sugar-specific proteins have been named lectins. Although shown to occur widely in plants and to some extent also in invertebrates, very few lectins had been isolated until the early 1970s, and they had attracted little attention. This attitude changed with the demonstration that lectins are extremely useful tools for the investigation of carbohydrates on cell surfaces, in particular of the changes that the latter undergo in malignancy, as well as for the isolation and characterization of glycoproteins. In subsequent years numerous lectins have been isolated from plants as well as from microorganisms and animals, and during the past two decades the structures of hundreds of them have been established. Concurrently, it was shown that lectins function as recognition molecules in cell-molecule and cell-cell interactions in a variety of biological systems. Here we present a brief account of 100-plus years of lectin research and show how these proteins have become the focus of intense interest for biologists and in particular for the glycobiologists among them.
Affinity Labels↗
Probing genetic variation and glycoform distribution in lectins of the Erythrina genus by mass spectrometry.
Six leguminous lectins from the seeds of plants of the Erythrina genus, namely E. caffra (ECafL), E. cristagalli (ECL), E. flabelliformis (EFL), E. lysistemon (ELysL), E. rubrinerva (ERL), and E. vespertilio (EVL), were examined to establish their sequence homology and to determine the structure and sites of attachment of their glycans. Tryptic digests of these lectins were analyzed by capillary electrophoresis coupled to electrospray mass spectrometry (CE-ESMS). Assignments were made by comparing the molecular masses of the observed tryptic peptides with those of Erythrina corallodendron lectin (ECorL), the sequence of which had been established previously. Glycan structure and genetic variations in the amino acid sequence were probed by tandem mass spectrometry. Small differences were found between the sequences of the various lectins examined and all of them exhibited C-terminal processing resulting in proteins with a C-terminal Asn residue. The major glycan of these glycoproteins was shown to be the heptasaccharide Man(3)XylFucGlcNAc(2), consistent with previous investigations on ECL and ECorL. A minor glycan heterogeneity was observed for most lectins examined except for that of ECafL and ECorL where an extra hexose residue was observed on the reducing GlcNAc residue of the heptasaccharide.
Amino Acid Sequence↗
A high molecular mass cranberry constituent reduces mutans streptococci level in saliva and inhibits in vitro adhesion to hydroxyapatite.
Previous investigations showed that a high molecular mass, non-dialyzable material (NDM) from cranberries inhibits the adhesion of a number of bacterial species and prevents the co-aggregation of many oral bacterial pairs. In the present study we determined the effect of mouthwash supplemented with NDM on oral hygiene. Following 6 weeks of daily usage of cranberry-containing mouthwash by an experimental group (n = 29), we found that salivary mutans streptococci count as well as the total bacterial count were reduced significantly (ANOVA, P < 0.01) compared with those of the control (n = 30) using placebo mouthwash. No change in the plaque and gingival indices was observed. In vitro, the cranberry constituent inhibited the adhesion of Streptococcus sobrinus to saliva-coated hydroxyapatite. The data suggest that the ability to reduce mutans streptococci counts in vivo is due to the anti-adhesion activity of the cranberry constituent.
Adult↗
Susceptibility of Helicobacter pylori isolates to the antiadhesion activity of a high-molecular-weight constituent of cranberry.
The sensitivity of a large number of antibiotic-resistant and nonresistant Helicobacter pylori isolates to the antiadhesion effect of a high-molecular-mass, nondialysable constituent of cranberry juice was tested. Confluent monolayers of gastric cell line in microtiter plate wells were exposed to bacterial suspensions prepared from 83 H. pylori isolates from antibiotic-treated and untreated patients in the presence and absence of the cranberry constituent. Urease assay was used to calculate the percentage of adhesion inhibition. In two thirds of the isolates, adhesion to the gastric cells was inhibited by 0.2 mg/mL of the nondialysable material. There was no relationship between the antiadhesion effect of the cranberry material and metronidazole resistance in isolates from either treated or untreated patients (N=35). Only 13 isolates (16%) were resistant to both the nondialysable material and metronidazole, and 30 (36%) were resistant to the nondialysable material alone. There was no cross-resistance to the nondialysable material and metronidazole. These data suggest that a combination of antibiotics and a cranberry preparation may improve H. pylori eradication.
Amoxicillin↗
Carbohydrates and glycoconjugates: progress in non-mammalian glycosylation, glycosyltransferases, invertebrate lectins and carbohydrate-carbohydrate interactions.
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Carbohydrate Metabolism↗
Letter to the Glyco-Forum: Karl Meyer, lysozyme, and penicilin.
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History, 20th Century↗
Role of N-linked glycan in the unfolding pathway of Erythrina corallodendron lectin.
Erythrina corallodendron lectin (ECorL) exhibits an exquisitely structured oligosaccharide chain. Interestingly, the bacterially expressed, nonglycosylated counterpart, rECorL, possesses an essentially identical carbohydrate specificity and agglutinating activity as the glycosylated lectin, thus suggesting that the overall structure of the two are identical. This paper reports the unfolding behavior of E. corallodendron lectin in its glycosylated (EcorL) and nonglycosylated (rECorL) forms. ECorL shows a two-state unfolding pattern during isothermal melts and differential scanning calorimetry (DSC). The T(g) of ECorL as obtained from isothermal melts is 74 degrees C at pH 7.4. The T(p) obtained from DSC studies is between 74.8 to 68.1 degrees C in the pH range of 5.26-7.77. The recombinant lectin (rECorL), which is devoid of carbohydrate, shows, in contrast to the glycosylated protein, a non-two-state unfolding profile as measured by both probes mentioned, but the number of intermediates during unfolding could not be ascertained. Simulated annealing on ECorL, with the sugars removed, reveals that the protein Calpha backbones overlap, indicating that the overall structure, including the mode of dimerization, of rECorL is insignificantly altered as compared to ECorL. The alterations in the folding behavior of rECorL as compared to that observed in ECorL may be due to the fact that, unlike most other glycoproteins, one of the glycans in ECorL is unusually structured and forms many hydrogen bonds with the protein. It therefore appears that while the covalently linked sugar does not contribute appreciably to the final folded structure of ECorL, it does alter its folding process in a significant manner.
Calcium↗
Anti-adhesion therapy of bacterial diseases: prospects and problems.
The alarming increase in drug-resistant bacteria makes a search for novel means of fighting bacterial infections imperative. An attractive approach is the use of agents that interfere with the ability of the bacteria to adhere to tissues of the host, since such adhesion is one of the initial stages of the infectious process. The validity of this approach has been unequivocally demonstrated in experiments performed in a wide variety of animals, from mice to monkeys, and recently also in humans. Here we review various approaches to anti-adhesion therapy, including the use of receptor and adhesin analogs, dietary constituents, sublethal concentrations of antibiotics and adhesin-based vaccines. Because anti-adhesive agents are not bactericidal, the propagation and spread of resistant strains is much less likely to occur than as a result of exposure to bactericidal agents, such as antibiotics. Anti-adhesive drugs, once developed, may, therefore, serve as a new means to fight infectious diseases.
Adhesins, Bacterial↗
Evolutionary analysis reveals collective properties and specificity in the C-type lectin and lectin-like domain superfamily.
Members of the C-type lectin/C-type lectin-like domain (CTL/CTLD) superfamily share a common fold and are involved in a variety of functions, such as generalized defense mechanisms against foreign agents, discrimination between healthy and pathogen-infected cells, and endocytosis and blood coagulation. In this work we used ConSurf, a computer program recently developed in our lab, to perform an evolutionary analysis of this superfamily in order to further identify characteristics of all or part of its members. Given a set of homologous proteins in the form of multiple sequence alignment (MSA) and an inferred phylogenetic tree, ConSurf calculates the conservation score in every alignment position, taking into account the relationships between the sequences and the physicochemical similarity between the amino acids. The scores are then color-coded onto the three-dimensional structure of one of the homologous proteins. We provide here and at http://ashtoret.tau.ac.il/ approximately sharon a detailed analysis of the conservation pattern obtained for the entire superfamily and for two subgroups of proteins: (a) 21 CTLs and (b) 11 heterodimeric CTLD toxins. We show that, in general, proteins of the superfamily have one face that is constructed mostly of conserved residues and another that is not, and we suggest that the former face is involved in binding to other proteins or domains. In the CTLs examined we detected a region of highly conserved residues, corresponding to the known calcium- and carbohydrate-binding site of the family, which is not conserved throughout the entire superfamily, and in the CTLD toxins we found a patch of highly conserved residues, corresponding to the known dimerization region of these proteins. Our analysis also detected patches of conserved residues with yet unknown function(s).
Amino Acid Sequence↗
Biases and complex patterns in the residues flanking protein N-glycosylation sites.
N-Glycosylation, the most common and most versatile protein modification reaction, occurs at the beta-amide of the aspargine of the Asn-Xaa-Ser/Thr sequon. For reasons that are unclear, not all such sequons are glycosylated. To find patterns that affect glycosylation, we examined the amino acid residues from the 20th preceding the sequon to the 20th residue following it, using bioinformatics tools. A clean data set of annotated, experimentally verified, glycosylated and nonglycosylated sequons derived from 617 well-defined nonredundant N- and N-,O-glycoproteins listed in SWISS-PROT (June 2002) was used. NXS and NXT sequons were analyzed separately. Although no overt patterns were found to explain sequon occupancy or nonoccupancy, trends for over- or underrepresentation of certain amino acids at particular positions were statistically significant and different in NXS and NXT sequons. In extension of earlier reports, none of the 80 Asn-Pro-Ser/Thr found were glycosylated, and a markedly low level of glycosylation was seen in sequons with Pro at the position following the Ser/Thr. In addition, a general observation was made that the considerable number of glycosylated sequons in the C-terminal 10 residues of glycoproteins suggests that N-glycosylation in these cases may be posttranslational and not cotranslational, as widely accepted.
Amino Acid Sequence↗
Recombinant plant lectins and their mutants.
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Amino Acid Sequence↗
How proteins bind carbohydrates: lessons from legume lectins.
The pioneering studies of Irvin Liener on soybean agglutinin (SBA) in the early 1950s served as the starting point of our involvement in lectin research during the past four decades. Initially we characterized SBA extensively as a glycoprotein and showed that its covalently linked glycan is an oligomannoside commonly present in animal glycoproteins. We have also introduced the use of the lectin to the study of normal and malignant cells and to the purging of bone marrow for transplantation. Our recent work focuses on the combining site of Erythrina corallodendron lectin, closely related to SBA. In this legume lectin, as in essentially all other members of the same protein family, irrespective of their sugar specificity, interactions with a constellation of three invariant residues (aspartic acid, asparagine, and an aromatic residue) are essential for ligand binding. Lectins from other families, whether of plants or animals, also combine with carbohydrates by H-bonds and hydrophobic interactions, but the amino acids involved may differ even if the specificity of the lectins is the same. Therefore, nature finds diverse solutions for the design of binding sites for structurally similar ligands, such as mono- or oligosaccharides. This diversity strongly suggests that lectins are products of convergent evolution.
Binding Sites↗
High-resolution crystal structures of Erythrina cristagalli lectin in complex with lactose and 2'-alpha-L-fucosyllactose and correlation with thermodynamic binding data.
The primary sequence of Erythrina cristagalli lectin (ECL) was mapped by mass spectrometry, and the crystal structures of the lectin in complex with lactose and 2'-alpha-L-fucosyllactose were determined at 1.6A and 1.7A resolution, respectively. The two complexes were compared with the crystal structure of the closely related Erythrina corallodendron lectin (ECorL) in complex with lactose, with the crystal structure of the Ulex europaeus lectin II in complex with 2'-alpha-L-fucosyllactose, and with two modeled complexes of ECorL with 2'-alpha-L-fucosyl-N-acetyllactosamine. The molecular models are very similar to the crystal structure of ECL in complex with 2'-alpha-L-fucosyllactose with respect to the overall mode of binding, with the L-fucose fitting snugly into the cavity surrounded by Tyr106, Tyr108, Trp135 and Pro134 adjoining the primary combining site of the lectin. Marked differences were however noted between the models and the experimental structure in the network of hydrogen bonds and hydrophobic interactions holding the L-fucose in the combining site of the lectin, pointing to limitations of the modeling approach. In addition to the structural characterization of the ECL complexes, an effort was undertaken to correlate the structural data with thermodynamic data obtained from microcalorimetry, revealing the importance of the water network in the lectin combining site for carbohydrate binding.
Amino Acid Sequence↗
Fighting infectious diseases with inhibitors of microbial adhesion to host tissues.
The majority of infectious diseases are initiated by the adhesion of pathogenic organisms to the tissues of the host. In many cases this adhesion is mediated by lectins present on the surface of the infectious organism that bind to complementary carbohydrates on the surface of the host tissues. Soluble carbohydrates recognized by the bacterial lectins block the adhesion of the bacteria to animal cells in vitro. Moreover, such carbohydrates have been shown to protect against experimental infection by lectin-carrying bacteria of different mammals, such as mice, rabbits, calves, and monkeys. Agents other than carbohydrates also block adhesion, as demonstrated with cranberry juice as well as with low and high molecular weight preparations isolated from the juice. Both kinds of preparation inhibited the adhesion in vitro of Escherichia coli to different animal cells. In addition, the high molecular weight material acted similarly on the adhesion of Helicobacter pylori to human gasrointenstinal cells, and on the coaggregation of oral bacteria. Furthermore, in limited clinical studies regular drinking of cranberry juice had a significant preventive effect on bacteriuria, and the high molecular weight constituent of the juices was also effective in decreasing the salivary level of Streptococus mutans, the major cause of tooth decay. Because the inhibitors of adhesion examined are not bactericidal, the selection of resistant inhibitor strains is unlikely to occur. Together, these findings may lead to new therapeutic strategies that are in dire need because of the world-wide increase in antibiotic resistant bacteria.
Animals↗