Biomedical subjects

E J Van Damme

Publications and source records attributed to E J Van Damme.

At least 19 recordsLinked to original sources

PubMed2001

Structure of a legume lectin from the bark of Robinia pseudoacacia and its complex with N-acetylgalactosamine.

The structure of the bark lectin RPbAI (isoform A4) from Robinia pseudoacacia has been determined by protein crystallography both in the free form and complexed with N-acetylgalactosamine. The free form is refined at 1.80 A resolution to an R-factor of 18.9% whereas the complexed structure has an R-factor of 19.7% at 2.05 A resolution. Both structures are compared to each other and to other available legume lectin structures. The polypeptide chains of the two structures exhibit the characteristic legume lectin tertiary fold. The quaternary structure resembles that of the Phaseolus vulgaris lectin, the soybean agglutinin, and the Dolichos biflorus lectin, but displays some unique features leading to the extreme stability of this lectin.

Acacia↗

PubMed2001

Type-1 ribosome-inactivating protein from iris (Iris hollandica var. Professor Blaauw) binds specific genomic DNA fragments.

The capacity of IRIP, a type-1 ribosome-inactivating protein (RIP) isolated from the bulbs of Iris hollandica, to bind specific DNA sequences from a mixture of approx. 200 bp (average length) fragments of total genomic DNA from Iris genome was studied. Fragments that were preferentially bound by IRIP were enriched by several cycles of affinity binding and PCR, and were cloned and sequenced. The selected DNA fragments do not share conserved sequences, indicating that IRIP does not bind DNA fragments in a strictly sequence-specific manner. According to sequence analysis, most IRIP-bound fragments contain one or more possible free energy-stable hairpin structure(s) in their secondary structure, which may be the basis for recognition between IRIP and these DNA fragments. Some, but not all, DNA fragments moderately lower the RNA N-glycosidase activity of IRIP towards rabbit reticulocyte lysate ribosomes. IRIP does not remove adenines from the binding fragments, which implies that it does not act as a polynucleotide:adenosine glycosidase towards these DNA fragments. The selective binding of IRIP to conspecific DNA fragments is also discussed in view of the novel concept that RIPs may act as DNA-binding proteins with a regulatory activity on gene expression.

Blotting, Southern↗

PubMed2001

The Gal/GalNAc-specific lectin from the plant pathogenic basidiomycete Rhizoctonia solani is a member of the ricin-B family.

The lectin isolated from the phytopathogenic basidiomycete Rhizoctonia solani (RSA) is a homodimer of two noncovalently associated monomers of 15.5 kDa. RSA is a basic protein (pI > 9) which consists mainly of beta-sheets. A presumed relationship with ricin-B is supported by the sequence similarity between the N-terminus of RSA and the N-terminal subdomain of ricin-B. Hydrophobic cluster analysis confirms that the N-terminus of both proteins has a comparable folding. RSA exhibits specificity towards Gal/GalNAc whereby the hydroxyls at the C3', C4', and C6' positions of the pyranose ring play a key role in the interaction with simple sugars. The carbohydrate-binding site of RSA apparently accommodates only a single sugar unit. Our results demonstrate an obvious evolutionary relationship between some fungal and plant lectins, but also provide evidence for the occurrence of a lectin consisting of subunits corresponding to a single subdomain of ricin-B.

Acetylgalactosamine↗

PubMed2001

Classification of plant lectins in families of structurally and evolutionary related proteins.

The majority of plant lectins can be classified in seven families of structurally and evolutionary related proteins. Within a given lectin family most but not necessarily all members are built up of protomers with a similar primary structure and overall 3-D fold. The overall structure of the native lectins is not only determined by the structure of the protomers but depends also on the degree of oligomerization and in some cases on the post-translational processing of the lectin precursors. In general, lectin families are fairly homogeneous for what concerns the overall specificity of the individual lectins, which illustrates that the 3-D structure of the binding site has been conserved during evolution. In the case of the jacalin-related lectins the occurrence of a mannose- and galactose-binding subfamily can be explained by the fact that a post-translational cleavage of the protomers (of the galactose-binding subfamily) yields a slightly altered binding site. Unlike the other families, the legume lectins display a wide range of specificites, which is clearly reflected in the occurrence of sugar-binding sites with a different 3-D structure.

Carbohydrate Metabolism↗

PubMed2001

Lectin histochemistry of microvascular endothelium in chick and quail musculature.

The lectin binding pattern of muscular microvessels in chick, quail and chick/quail chimeras was analysed. Paraffin wax sections of muscles from embryonic and adult animals were used. The biotin-labelled lectins were detected by avidin-alkaline phosphatase complex. The following lectins bound to muscular microvessels including arterioles, capillaries and venules of both species: SNA-I (Sambucus nigra agglutinin), MAA (Maackia amurensis agglutinin), AIA (Artocarpus integrifolia agglutinin), VAA-I, VAA-II and VAA-III (Viscum album agglutinin I-III), WGA (wheat germ agglutinin), LEA (Lycopersicon esculentum agglutinin). Endomysium and basement membranes of muscle fibres were also stained to a variable extent and intensity. Only SNA-I stained almost exclusively the endothelium of blood vessels. WFA (Wisteria floribunda agglutinin) bound to the quail endothelium only. MPA (Maclura pomifera agglutinin) marked vessels in adult muscles of chick and quail, but embryonic vessels were stained in quail only. Our results show that lectin histochemistry is a useful tool for visualisation of microvasculature in avian species. In particular, WFA and MPA can be used to determine the origin of endothelia in chick/quail chimeras.

Animals↗

PubMed2001

Mannose-binding plant lectins: different structural scaffolds for a common sugar-recognition process.

Mannose-specific lectins are widely distributed in higher plants and are believed to play a role in recognition of high-mannose type glycans of foreign micro-organisms or plant predators. Structural studies have demonstrated that the mannose-binding specificity of lectins is mediated by distinct structural scaffolds. The mannose/glucose-specific legume (e.g., Con A, pea lectin) exhibit the canonical twelve-stranded beta-sandwich structure. In contrast to legume lectins that interact with both mannose and glucose, the monocot mannose-binding lectins (e.g., the Galanthus nivalis agglutinin or GNA from bulbs) react exclusively with mannose and mannose-containing N-glycans. These lectins possess a beta-prism structure. More recently, an increasing number of mannose-specific lectins structurally related to jacalin (e.g., the lectins from the Jerusalem artichoke, banana or rice), which also exhibit a beta-prism organization, were characterized. Jacalin itself was re-defined as a polyspecific lectin which, in addition to galactose, also interacts with mannose and mannose-containing glycans. Finally the B-chain of the type II RIP of iris, which has the same beta-prism structure as all other members of the ricin-B family, interacts specifically with mannose and galactose. This structural diversity associated with the specific recognition of high-mannose type glycans highlights the importance of mannose-specific lectins as recognition molecules in higher plants.

Binding Sites↗

PubMed2001

Lectin and proteoglycan histochemistry of Merkel cell carcinomas.

Changes in carbohydrate residue expression and in proteoglycan distribution occur during different stages of tumor development and progression. However, few data concerning carbohydrate residue analysis as performed by lectin histochemistry and proteoglycan distribution of Merkel cell carcinoma, a rare malignant tumor of the skin, have been reported. Hence, lectin- and proteoglycan immunohistochemistry was performed on paraffin wax material of 9 cases of Merkel cell carcinomas characterized by cytokeratin and neurofilament immunohistochemistry. The lectin binding pattern of tumor cells varied between lectins with different sugar binding specificities, while within a given nominal sugar specificity intensities were remarkably similar between tumors from different patients. The most intensive reaction was observed using Con A (mannose/glucose-specific) followed by LCA with the same specificity and the N-Acetyl glucosamine-specific lectins (WGA, UDA, CMA), while no fucose binding sites were detected (UEA-I). In addition, N-Acetyl galactosamine residues were only occasionally detected. The lectin binding pattern of Merkel cell carcinoma cells indicated that predominantly N-linked glycans and not O-linked glycans, typical for mucins of most epithelia, were present. Hence these tumor cells were relatively undifferentiated and resembled stem cells more closely than differentiated epithelia. The tumor stroma was especially evaluated in this study and showed a lectin reaction, which was intermediate between the tumor cells and extra-tumoral stroma. For example, the reactions of N-Acetyl galactosamine-specific lectins were intensive in the extra-tumoral stroma but nearly negative in tumor cells, while the lectin reaction of the intra-tumoral stroma was similar to the cellular reaction. These results indicated an influence of tumor cells on the stromal constituents. Antibodies against chondroitin type glycosaminoglycans reacted with the tumor stroma and the pericellular substance around the tumor cells most intensely in - and around the major tumor septae which, in general, were well vascularized. The most intensive immunoreactivity was detected using the chondroitin-6-sulfate antibody. The cellular and membrane-associated reaction for heparan sulfate was less intensive in comparison to epidermal cells. In conclusion the pattern of lectin-binding sites, the high chondroitin(sulfate) specific reactivity and the relatively low intensity of heparan sulfate immunohistochemistry indicate a low degree of differentiation and high malignity of the tumors, which is consistent with the clinical behavior of Merkel cell carcinomas.

Biomarkers, Tumor↗

PubMed2001

Purification, characterization, immunolocalization and structural analysis of the abundant cytoplasmic beta-amylase from Calystegia sepium (hedge bindweed) rhizomes.

An abundant catalytically active beta-amylase (EC 3.2.1.2) was isolated from resting rhizomes of hedge bindweed (Calystegia sepium). Biochemical analysis of the purified protein, molecular modeling, and cloning of the corresponding gene indicated that this enzyme resembles previously characterized plant beta-amylases with regard to its amino-acid sequence, molecular structure and catalytic activities. Immunolocalization demonstrated that the beta-amylase is exclusively located in the cytoplasm. It is suggested that the hedge bindweed rhizome beta-amylase is a cytoplasmic vegetative storage protein.

Amino Acid Sequence↗

PubMed2001

Gastrodianin-like mannose-binding proteins: a novel class of plant proteins with antifungal properties.

The orchid Gastrodia elata depends on the fungus Armillaria mellea to complete its life cycle. In the interaction, fungal hyphae penetrate older, nutritive corms but not newly formed corms. From these corms, a protein fraction with in vitro activity against plant-pathogenic fungi has previously been purified. Here, the sequence of gastrodianin, the main constituent of the antifungal fraction, is reported. Four isoforms that encoded two different mature proteins were identified at the cDNA level. Another isoform was detected in sequenced peptides. Because the antifungal activity of gastrodianins produced in and purified from Escherichia coli and Nicotiana tabacum was comparable to that of gastrodianin purified from the orchid, gastrodianins are the active component of the antifungal fractions. Gastrodianin accumulation is probably an important part of the mechanism by which the orchid controls Armillaria penetration. Gastrodianin was found to be homologous to monomeric mannose-binding proteins of other orchids, of which at least one (Epipactis helleborine mannose-binding protein) also displayed in vitro antifungal activity. This establishes the gastrodianin-like proteins (GLIPs) as a novel class of antifungal proteins.

Amino Acid Sequence↗

PubMed2001

Carbohydrate binding properties of banana (Musa acuminata) lectin I. Novel recognition of internal alpha1,3-linked glucosyl residues.

Examination of lectins of banana (Musa acuminata) and the closely related plantain (Musa spp.) by the techniques of quantitative precipitation, hapten inhibition of precipitation, and isothermal titration calorimetry showed that they are mannose/glucose binding proteins with a preference for the alpha-anomeric form of these sugars. Both generate precipitin curves with branched chain alpha-mannans (yeast mannans) and alpha-glucans (glycogens, dextrans, and starches), but not with linear alpha-glucans containing only alpha1,4- and alpha1,6-glucosidic bonds (isolichenan and pullulan). The novel observation was made that banana and plantain lectins recognize internal alpha1,3-linked glucosyl residues, which occur in the linear polysaccharides elsinan and nigeran. Concanavalin A and lectins from pea and lentil, also mannose/glucose binding lectins, did not precipitate with any of these linear alpha-glucans. This is, the authors believe, the first report of the recognition of internal alpha1,3-glucosidic bonds by a plant lectin. It is possible that these lectins are present in the pulp of their respective fruit, complexed with starch.

Binding Sites↗

PubMed2001

Carbohydrate binding properties of banana (Musa acuminata) lectin II. Binding of laminaribiose oligosaccharides and beta-glucans containing beta1,6-glucosyl end groups.

This paper extends our knowledge of the rather bizarre carbohydrate binding poperties of the banana lectin (Musa acuminata). Although a glucose/mannose binding protein which recognizes alpha-linked gluco-and manno-pyranosyl groups of polysaccharide chain ends, the banana lectin was shown to bind to internal 3-O-alpha-D-glucopyranosyl units. Now we report that this lectin also binds to the reducing glucosyl groups of beta-1,3-linked glucosyl oligosaccharides (e.g. laminaribiose oligomers). Additionally, banana lectin also recognizes beta1,6-linked glucosyl end groups (gentiobiosyl groups) as occur in many fungal beta1,3/1,6-linked polysaccharides. This behavior clearly distinguishes the banana lectin from other mannose/glucose binding lectins, such as concanavalin A and the pea, lentil and Calystegia sepium lectins.

Binding Sites↗

PubMed2001

Preparation of monospecific polyclonal antibodies against Sambucus nigra lectin related protein, a glycosylated plant protein.

A simple, but highly efficient, method was developed for the purification of monospecific antibodies against the plant glycoprotein Sambucus nigra lectin related protein. In a first step, the antiserum is purified by affinity chromatography on a column with the immobilized antigen. To deplete the affinity-purified antiserum from aspecific cross-reacting antibodies directed against the glycan part of the glycoprotein, a second affinity chromatography on an unrelated plant glycoprotein, in casu the Robinia pseudoacacia agglutinin, is included.

Animals↗

PubMed2001

Ribosome-inactivating proteins from plants: more than RNA N-glycosidases?

Many plants contain proteins that are capable of inactivating ribosomes and accordingly are called ribosome-inactivating proteins or RIPs. These typical plant proteins receive a lot of attention in biological and biomedical research because of their unique biological activities toward animal and human cells. In addition, evidence is accumulating that some RIPs play a role in plant defense and hence can be exploited in plant protection. To understand the mode of action of RIPs and to optimize their medical and therapeutical applications and their use as antiviral compounds in plant protection, intensive efforts have been made to unravel the enzymatic activities of RIPs and provide a structural basis for these activities. Though marked progress has been made during the last decade, the enzymatic activity of RIPs has become a controversial issue because of the concept that RIPs possess, in addition to their classical RNA N-glycosidase and polynucleotide:adenosine glycosidase activity, other unrelated enzymatic activities. Moreover, the presumed novel enzymatic activities, especially those related to diverse nuclease activities, are believed to play an important role in various biological activities of RIPs. However, both the novel enzymatic activities and their presumed involvement in the biological activities of RIPs have been questioned because there is evidence that the activities observed are due to contaminating enzymes. We offer a critical review of the pros and cons of the putative novel enzymatic activities of RIPs. Based on the available data, it is suggested that there is little conclusive evidence in support of the presumed activities and that in the past too little attention has been given to the purity of the RIP preparation. The antiviral activity and mode of action of RIPs in plants are discussed in view of their classical and presumed novel enzymatic activities.

Antiviral Agents↗

PubMed2001

Iris bulbs express type 1 and type 2 ribosome-inactivating proteins with unusual properties.

Two closely related lectins from bulbs of the Dutch iris (Iris hollandica var. Professor Blaauw) have been isolated and cloned. Both lectins, called Iris agglutinin b and Iris agglutinin r, possess N-glycosidase activity and share a high sequence similarity with previously described type 2 ribosome-inactivating proteins (RIP). However, these lectins show only 57% to 59% sequence identity to a previously characterized type 1 RIP from iris, called IRIP. The identification of the iris lectins as type 2 RIP provides unequivocal evidence for the simultaneous occurrence of type 1 and type 2 RIP in iris bulbs and allowed a detailed comparison of type 1 and type 2 RIP from a single plant, which provides further insight into the molecular evolution of RIP. Binding studies and docking experiments revealed that the lectins exhibit binding activity not only toward Gal/N-acetylgalactosamine, but also toward mannose, demonstrating for the first time that RIP-binding sites can accommodate mannose.

Amino Acid Sequence↗

PubMed2001

The crystals of a mannose-specific jacalin-related lectin from Morus nigra are merohedrally twinned.

MornigaM, a lectin from Morus nigra, belongs to the mannose-binding subgroup of the family of jacalin-related plant lectins. It was crystallized in the P6(5) space group, with unit-cell parameters a = b = 110.74, c = 159.28 A. The partially merohedrally twinned crystals could be detwinned and a subsequent molecular-replacement solution could be found using the coordinates of jacalin. Preliminary analysis clearly shows the tetrameric assembly of this protein. Furthermore, data from MornigaM crystals soaked in a mannose solution were collected.

Crystallization↗

PubMed2000

Microenvironment of cysteine 242 in type-1 ribosome-inactivating protein from iris.

IRIP is a type-1 ribosome-inactivating protein isolated from the bulbs of Iris hollandica. It is one of the few type-1 RIPs that contain Cys residue(s) in their primary sequence. IRIP contains a single Cys residue at position 242. Although IRIP is thought to be a monomeric protein, SDS-PAGE indicates that part of the IRIP molecules can exist as disulphide bridge-linked dimers. Probing of the reactivity of the unique Cys residue by 5, 5'-dithiobis(2-nitrobenzoic acid) indicates that Cys(242) in IRIP is free but is only partially accessible to modifiers. Molecular modelling of IRIP is in agreement with this conclusion. Binding of the ligands adenine and poly(A) results in little or no effect on the conformation of Cys(242) in IRIP. Chemical modification of IRIP by a specific thiol modifier does not abolish the RNA N-glycosidase activity of IRIP, suggesting that Cys(242) is not critical for the enzymatic activity of IRIP. These results suggest that IRIP has the potential to be developed as a novel immunotoxin.

Amino Acid Sequence↗

PubMed2000

Accessibility of the high-mannose glycans of glycoprotein gp120 from human immunodeficiency virus type 1 probed by in vitro interaction with mannose-binding lectins.

The direct interaction of mannose-specific plant lectins with gp120 of HIV-1 was studied by surface plasmon resonance. Inhibition experiments indicated that exposed high mannose type glycans play a key role in the interaction. Most of the lectins specifically accommodate outer alpha1,2-, alpha1,3-, or alpha1,6-linked di- or trimannosides, and especially legume lectins, also interact with the trimannoside core of the complex type glycans. The unexpected affinity of some lectins towards gp120 presumably results from conformational differences in their binding sites. These results demonstrate that mannose-specific plant lectins are powerful tools to study the accessibility and elucidate the function of the gp120 glycans in the recognition and infection of the host cells by HIV-1.

Binding Sites↗

PubMed2000

The galactose-binding and mannose-binding jacalin-related lectins are located in different sub-cellular compartments.

A galactose-specific and a mannose-specific lectin of the family of the jacalin-related lectins have been localized by immunofluorescence microscopy. The present localization studies provide for the first time unambiguous evidence for the cytoplasmic location of the mannose-specific jacalin-related lectin from rhizomes of Calystegia sepium, which definitely differs from the vacuolar location of the galactose-specific jacalin from Artocarpus integrifolia. These observations support the hypothesis that the galactose-specific jacalin-related lectins evolved from their mannose-specific homologues through the acquisition of vacuolar targeting sequences.

Cell Compartmentation↗