Related Lectins Research Articles

Interactions of LSECtin and DC-SIGN/DC-SIGNR with viral ligands: Differential pH dependence, internalization and virion binding

The calcium-dependent lectins DC-SIGN and DC-SIGNR (collectively termed DC-SIGN/R) bind to high-mannose carbohydrates on a variety of viruses. In contrast, the related lectin LSECtin does not recognize mannose-rich glycans and interacts with a more restricted spectrum of viruses. Here, we analyzed whether these lectins differ in their mode of ligand engagement. LSECtin and DC-SIGNR, which we found to be co-expressed by liver, lymph node and bone marrow sinusoidal endothelial cells, bound to soluble Ebola virus glycoprotein (EBOV-GP) with comparable affinities. Similarly, LSECtin, DC-SIGN and the Langerhans cell-specific lectin Langerin readily bound to soluble human immunodeficiency virus type-1 (HIV-1) GP. However, only DC-SIGN captured HIV-1 particles, indicating that binding to soluble GP is not necessarily predictive of binding to virion-associated GP. Capture of EBOV-GP by LSECtin triggered ligand internalization, suggesting that LSECtin like DC-SIGN might function as an antigen uptake receptor. However, the intracellular fate of lectin–ligand complexes might differ. Thus, exposure to low-pH medium, which mimics the acidic luminal environment in endosomes/lysosomes, released ligand bound to DC-SIGN/R but had no effect on LSECtin interactions with ligand. Our results reveal important differences between pathogen capture by DC-SIGN/R and LSECtin and hint towards different biological functions of these lectins.

The mistletoe lectin I—Phloretamide structure reveals a new function of plant lectins

The X-ray structure at 2.7 Å resolution of the complex between the European mistletoe lectin I ( Viscum album, ML-I) and the plant growth hormone, 3-( p-hydroxyphenyl)-propionic acid amide (phloretamide, PA) from xylem sap has revealed the binding of PA at the so far undescribed hydrophobic cavity located between the two subunits of this ribosome-inhibiting protein. No such cavity is observed in related lectins. The binding of PA is achieved through interactions with the non-conserved residues Val228A, Leu230A, Arg388B, and the C-terminal Pro510B. It is conceivable that binding of PA to ML-I is part of a defence mechanism of the parasite against the host, whereby the parasite prevents the growth hormone of the host from interfering with its own regulatory system. The specific binding of PA to ML-I indicates that heterodimeric RIPs are multifunctional proteins whose functions in the cell have not yet been fully recognized and analyzed.

The DC-SIGN–related lectin LSECtin mediates antigen capture and pathogen binding by human myeloid cells

AbstractLiver and lymph node sinusoidal endothelial cell C-type lectin (LSECtin [CLEC4G]) is a C-type lectin encoded within the liver/lymph node–specific intercellular adhesion molecule-3–grabbing nonintegrin (L-SIGN)/dendritic cell–specific intercellular adhesion molecule-3–grabbing nonintegrin (DC-SIGN)/CD23 gene cluster. LSECtin expression has been previously described as restricted to sinusoidal endothelial cells of the liver and lymph node. We now report LSECtin expression in human peripheral blood and thymic dendritic cells isolated ex vivo. LSECtin is also detected in monocyte-derived macrophages and dendritic cells at the RNA and protein level. In vitro, interleukin-4 (IL-4) induces the expression of 3 LSECtin alternatively spliced isoforms, including a potentially soluble form (Δ2 isoform) and a shorter version of the prototypic molecule (Δ3/4 isoform). LSECtin functions as a pathogen receptor, because its expression confers Ebola virus–binding capacity to leukemic cells. Sugar-binding studies indicate that LSECtin specifically recognizes N-acetyl-glucosamine, whereas no LSECtin binding to Mannan- or N-acetyl-galactosamine–containing matrices are observed. Antibody or ligand-mediated engagement triggers a rapid internalization of LSECtin,which is dependent on tyrosine and diglutamic-containing motifs within the cytoplasmic tail. Therefore, LSECtin is a pathogen-associated molecular pattern receptor in human myeloid cells. In addition, our results suggest that LSECtin participates in antigen uptake and internalization, and might be a suitable target molecule in vaccination strategies.

Complement protein C1q induces maturation of human dendritic cells

Maturation of dendritic cells (DCs) is known to be induced by several stimuli, including microbial products, inflammatory cytokines and immobilized IgG, as demonstrated recently. Since immune complexes formed in vivo also contain C1q, moreover apoptotic cells and several pathogens fix C1q in the absence of antibodies, we undertook to investigate whether this complement protein has an impact on various functions of human DCs. Maturation of monocyte-derived immature DCs (imMDCs) cultured on immobilized C1q was followed by monitoring expression of CD80, CD83, CD86, MHCII and CCR7. The functional activity of the cells was assessed by measuring cytokine secretion and their ability to activate allogeneic T lymphocytes. Cytokine production by T cells co-cultured with C1q-matured DCs was also investigated. C1q, but not the structurally related mannose-binding lectin was found to bind to imMDC in a dose-dependent manner and induced NF-κB translocation to the nucleus. Immobilized C1q induced maturation of MDCs and enhanced secretion of IL-12 and TNF-α, moreover, elevated their T-cell stimulating capacity. As IFN-γ levels were increased in supernatants of MDC–T cell co-cultures, our data suggest that C1q-induced DC maturation generates a Th1-type response. Interestingly, IL-10 levels were elevated by C1q-treated MDCs but not in the supernatant of their co-cultures with allogeneic T cells. Taken together, these results indicate that C1q-opsonized antigens may play a role in the induction and regulation of immune response. Moreover our data are relevant in view of the role of C1q in removal of apoptotic cells and the association between C1q-deficiency and autoimmunity.

The three-dimensional structure of codakine and related marine C-type lectins

Codakine is a new Ca 2+-dependent mannose-binding C-type lectin (MBL) isolated from the gill tissue of the tropical clam, Codakia orbicularis. Bioinformatic analyses with the BLAST program have revealed similarities with marine lectins involved in immunity whose three-dimensional (3D) structures were unknown up until recently. In this article, we present bioinformatic analyses of marine lectins that are homologous to codakine, in particular lectins from the sea worm Laxus oneistus, named mermaid. These lectins are involved in the symbiotic association with sulphur-oxidizing bacteria which are closely related to the C. orbicularis gill symbiont. Using homology modelling, folding that is characteristic of C-type lectins was observed in all the marine Ca 2+-dependent lectins studied, with conservation of random coiled structures of the carbohydrate recognition domain (CRD) and Ca 2+-binding sites. Like codakine, the marine lectins analysed contain a signal peptide commonly found in secreted and transmembrane proteins. The majority of the predictive 3D models established from the lectins exhibit a common feature, namely the involvement in invertebrate and vertebrate immunity (dendritic cell receptor, macrophage receptor, etc.). These bioinformatic analyses and the literature data support the hypothesis that codakine, like the L. oneistus mermaids, is probably involved in the cellular mediation of symbiosis and defence against pathogenic microorganisms.

The Crystal Structure and Mutational Binding Analysis of the Extracellular Domain of the Platelet-activating Receptor CLEC-2

The human C-type lectin-like molecule CLEC-2 is expressed on the surface of platelets and signaling through CLEC-2 causes platelet activation and aggregation. CLEC-2 is a receptor for the platelet-aggregating snake venom protein rhodocytin. It is also a newly identified co-receptor for human immunodeficiency virus type 1 (HIV-1). An endogenous ligand has not yet been identified. We have solved the crystal structure of the extracellular domain of CLEC-2 to 1.6-A resolution, and identified the key structural features involved in ligand binding. A semi-helical loop region and flanking residues dominate the surface that is available for ligand binding. The precise distribution of hydrophobic and electrostatic features in this loop will determine the nature of any endogenous ligand with which it can interact. Major ligand-induced conformational change in CLEC-2 is unlikely as its overall fold is compact and robust. However, ligand binding could induce a tilt of a 3-10 helical portion of the long loop region. Mutational analysis and surface plasmon resonance binding studies support these observations. This study provides a framework for understanding the effects of rhodocytin venom binding on CLEC-2 and for understanding the nature of likely endogenous ligands and will provide a basis for rational design of drugs to block ligand binding.

Plasma Levels of Mannan‐Binding Lectin in Relation to Periodontitis and Smoking

Mannan-binding lectin (MBL) is an important molecule of innate immunity; it acts as an opsonin and stimulates the classical complement pathway. Moreover, it has been suggested that MBL acts as a weak acute phase protein. We investigated whether MBL levels are increased in periodontitis, and we tested whether individuals deficient for MBL are more susceptible to periodontitis. A total of 219 subjects participated in the study. Plasma samples from 115 periodontitis patients and 104 healthy controls were taken, and the MBL levels were measured by enzyme-linked immunosorbent assay (ELISA). MBL levels were analyzed in relation to periodontitis, taking into consideration age, gender, ethnic and educational background, and smoking status. In some analyses, subjects with MBL plasma levels <0.8 microg/ml were considered MBL deficient. MBL plasma concentrations were not significantly different in moderate and severe periodontitis compared to controls (1.6, 1.4, and 1.6 microg/ml, respectively). Also, the prevalence of MBL deficiency was not found to be different between controls and moderate and severe periodontitis (45%, 37%, and 36%). However, among all subjects and among the non-deficient subjects, MBL levels were markedly increased in heavy smokers (>10 cigarettes per day), irrespective of periodontal disease status, in comparison to non-smokers and light smokers. MBL plasma levels did not show a correlation with plasma C-reactive protein (CRP) and were also not related to the prevalence of specific periodontal pathogens. MBL levels were not elevated in periodontitis, and MBL deficiency was not related to susceptibility for periodontitis. The fact that MBL levels were higher among heavy smokers is the subject of further investigation.

Comparative genetics and innate immune functions of collagenous lectins in animals

Collagenous lectins such as mannan-binding lectins (MBLs), ficolins (FCNs), surfactant proteins A and D (SP-A, SP-D), conglutinin (CG), and related ruminant lectins are multimeric proteins with carbohydrate-binding domains aligned in a manner that facilitates binding to microbial surface polysaccharides. MBLs and FCNs are structurally related to C1q, but activate the lectin complement pathway via interaction with MBL-associated serine proteases (MASPs). MBLs, FCNs, and other collagenous lectins also bind to some host macromolecules and contribute to their removal. While there is evidence that some lectins and the lectin complement pathway are conserved in vertebrates, many differences in collagenous lectins have been observed among humans, rodents, and other vertebrates. For example, humans have only one MBL but three FCNs, whereas most other species express two FCNs and two MBLs. Bovidae express CG and other SP-D-related collectins that are not found in monogastric species. Some dysfunctions of human MBL are due to single nucleotide polymorphisms (SNPs) that affect its expression or structure and thereby increase susceptibility to some infections. Collagenous lectins have well-established roles in innate immunity to various microorganisms, so it is possible that some lectin genotypes or induced phenotypes influence resistance to some infectious or inflammatory diseases in animals.

Childhood Levels of Immunoglobulins and Mannan‐Binding Lectin in Relation to Infections and Allergy

Respiratory tract infections, allergies and otitis media are common problems in early childhood. Our aim was to evaluate in a longitudinal community-based cohort study the association between maturation of immunoglobulin (Ig) and mannan-binding lectin (MBL) responses and disease manifestations in the first 4 years of life. Sustained low levels of IgA proved the strongest single indicator of susceptibility for recurrent otitis media (P = 0.008) and respiratory tract infections (P = 0.02), and this condition was also associated with low production of IgG subclasses. About 7% of the cohort had sustained low levels of MBL (<0.4 mg/l). Low MBL did not predispose to any ailments studied, but children with low IgA and recurrent otitis media had relatively low MBL at birth, which failed to increase during the study period and was significantly reduced at the age of 4 years (P = 0.04). MBL levels increased from birth to 2 years (P < 0.0001) and were higher in children than in adults (P = 0.001). The increase was 1.9-fold in children with no recorded clinical events and 1.7-fold in children with asthma or infections, but significantly lower, 1.2-fold, in children with recurrent otitis media. Low levels of IgA within the normal range may reveal disease susceptibility not detected by conventional criteria. Slow maturation of Ig appears to be the main factor of susceptibility during childhood, but a strong corollary role for MBL is indicated by the high levels produced during childhood as well as the precipitation of disease in children with low levels of MBL and Ig.

Marked Depletion of Glycosylation Sites in HIV-1 gp120 under Selection Pressure by the Mannose-Specific Plant Lectins ofHippeastrumHybrid andGalanthus nivalis

The plant lectins from Hippeastrum hybrid (HHA) and Galanthus nivalis (GNA) are 50,000-D tetramers showing specificity for alpha-(1,3) and/or alpha-(1,6)-mannose oligomers. They inhibit HIV-1 infection at a 50% effective concentration of 0.2 to 0.3 microg/ml. Escalating HHA or GNA concentrations (up to 500 microg/ml) led to the isolation of three HIV-1(III(B)) strains in CEM T cell cultures that were highly resistant to HHA and GNA, several other related mannose-specific plant lectins, and the monoclonal antibody 2G12, modestly resistant to the mannose-specific cyanovirin, which is derived from a blue-green alga, but fully susceptible to other HIV entry inhibitors as well as HIV reverse transcriptase inhibitors. These mutant virus strains were devoid of up to seven or eight of 22 glycosylation sites in the viral envelope glycoprotein gp120 because of mutations at the Asn or Thr/Ser sites of the N-glycosylation motifs. In one of the strains, a novel glycosylation site was created near a deleted glycosylation site. The affected glycosylation sites were predominantly clustered in regions of gp120 that are not involved in the direct interaction with either CD4, CCR5, CXCR4, or gp41. The mutant viruses containing the deleted glycosylation sites were markedly more infectious in CEM T-cell cultures than wild-type virus.

Isolation and characterization of a new mannose-binding lectin gene fromTaxus media

In this paper, we report the cloning and characterization of the first mannose-binding lectin gene from a gymnosperm plant species, Taxus media. The full-length cDNA of T. media agglutinin (TMA) consisted of 676 bp and contained a 432 bp open reading frame (ORF) encoding a 144 amino acid protein. Comparative analysis showed that TMA had high homology with many previously reported plant mannose-binding lectins and that tma encoded a precursor lectin with a 26-aa signal peptide. Molecular modelling revealed that TMA was a new mannosebinding lectin with three typical mannose-binding boxes like lectins from species of angiosperms. Tissue expression pattern analyses revealed that tma is expressed in a tissue-specific manner in leaves and stems, but not in fruits and roots. Phylogenetic tree analyses showed that TMA belonged to the structurally and evolutionarily closely related monocot mannose-binding lectin superfamily. This study provides useful information to understand the molecular evolution of plant lectins.

Study of real-time lectin–carbohydrate interactions on the surface of a quartz crystal microbalance

A quartz crystal microbalance (QCM) biosensor system for lectin-carbohydrate interactions has been developed. Yeast mannan was immobilised on polystyrene-coated quartz crystals, and interactions tested with the lectin concanavalin A (Con A). The biosensor could be easily operated, where mannan immobilisation and all binding analyses were performed in real-time using a flow-through system. The apparent binding constant for yeast mannan to Con A was estimated to be 0.4 microM, well in accordance to reported literature values. In addition, the effective concentration values (EC50-values) for a series of mannose/mannoside ligands, acting as competitors to the mannan/Con A interaction, were determined to range from 0.18 to 5.3 mM, in good correlation with a related enzyme-labelled lectin assay (ELLA) protocol.

DC-SIGN and L-SIGN Are High Affinity Binding Receptors for Hepatitis C Virus Glycoprotein E2

The hepatitis C virus (HCV) genome codes for highly mannosylated envelope proteins, which are naturally retained in the endoplasmic reticulum. We found that the HCV envelope glycoprotein E2 binds the dendritic cell-specific intercellular adhesion molecule 3-grabbing nonintegrin (DC-SIGN) and the related liver endothelial cell lectin L-SIGN through high-mannose N-glycans. Competing ligands such as mannan and an antibody directed against the carbohydrate recognition domains (CRD) abrogated binding. While no E2 interaction with distant monomeric CRDs on biosensor chips could be detected, binding is observed if CRDs are closely seeded (Kd = 48 nm) and if the CRD is part of the oligomeric-soluble extracellular domain of DC-SIGN (Kd = 30 nm). The highest affinity is seen for plasma membrane-expressed DC-SIGN and L-SIGN (Kd = 3 and 6 nm, respectively). These results indicate that several high-mannose N-glycans in a structurally defined cluster on E2 bind to several subunits of the oligomeric lectin CRD. High affinity interaction of viral glycoproteins with oligomeric lectins might represent a strategy by which HCV targets to and concentrates in the liver and infects dendritic cells.

Hepatitis C virus glycoproteins interact with DC-SIGN and DC-SIGNR.

DC-SIGN and DC-SIGNR are two closely related membrane-associated C-type lectins that bind human immunodeficiency virus (HIV) envelope glycoprotein with high affinity. Binding of HIV to cells expressing DC-SIGN or DC-SIGNR can enhance the efficiency of infection of cells coexpressing the specific HIV receptors. DC-SIGN is expressed on some dendritic cells, while DC-SIGNR is localized to certain endothelial cell populations, including hepatic sinusoidal endothelial cells. We found that soluble versions of the hepatitis C virus (HCV) E2 glycoprotein and retrovirus pseudotypes expressing chimeric forms of both HCV E1 and E2 glycoproteins bound efficiently to DC-SIGN and DC-SIGNR expressed on cell lines and primary human endothelial cells but not to other C-type lectins tested. Soluble E2 bound to immature and mature human monocyte-derived dendritic cells (MDDCs). Binding of E2 to immature MDDCs was dependent on DC-SIGN interactions, while binding to mature MDDCs was partly independent of DC-SIGN, suggesting that other cell surface molecules may mediate HCV glycoprotein interactions. HCV interactions with DC-SIGN and DC-SIGNR may contribute to the establishment or persistence of infection both by the capture and delivery of virus to the liver and by modulating dendritic cell function.

DC-SIGN (dendritic cell-specific ICAM-grabbing non-integrin) and DC-SIGN-related (DC-SIGNR): friend or foe?

C-type lectins are calcium-dependent carbohydrate-binding proteins with a wide range of biological functions, many of which are related to immunity. DC-SIGN (dendritic cell-specific ICAM-grabbing non-integrin, where ICAM is intercellular adhesion molecule) is a recently described mannose-specific C-type lectin expressed by dendritic cells. Dendritic cells are potent antigen-presenting cells capable of activating T-lymphocytes. DC-SIGN, which is expressed by dendritic cells, binds to ICAM-3 on T-lymphocytes, therefore playing an important role in the activation of T-lymphocytes. DC-SIGN can also bind HIV, and the virus may remain bound to DC-SIGN for protracted periods. DC-SIGN may deliver bound HIV to permissive cell types, mediating infection with high efficiency. A closely related C-type lectin, DC-SIGN-related molecule (DC-SIGNR) has also been described. DC-SIGNR is expressed by restricted subsets of endothelial cells, but has similar ICAM-3 and HIV-binding properties to DC-SIGN. This review describes the mapping of DC-SIGN and DC-SIGNR to chromosome 19p13.3 adjacent to the previously described C-type lectin, CD23 [the low-affinity receptor for immunoglobulin E (FcERII)]. The similar genomic organization of these three genes is discussed and consideration is given to the evolutionary duplications that may underlie this arrangement. Both DC-SIGN and DC-SIGNR possess a neck region, made up of multiple repeats, which supports the ligand-binding domain. Consideration is given to the biological reasons underlying the considerable polymorphism in the numbers of repeats in DC-SIGNR, but not DC-SIGN. The expression patterns of both DC-SIGN and DC-SIGNR are discussed in detail, with particular attention to the expression of both molecules in the placenta, which may have implications for the vertical transmission of HIV. Since dendritic cells may be important in determining the phenotype of many immune responses, via effects on T-lymphocytes, the differential expression of DC-SIGN by particular dendritic cell subsets may have important implications for the immunobiological functions of DC-SIGN. Similarly, the expression of DC-SIGNR by very restricted subsets of endothelial cells may give clues to the function of DC-SIGNR. Finally, the role of DC-SIGN in pathology, particularly in infective and neoplastic processes, is discussed, followed by speculation about likely future developments in this field.

Lectin domains on cytokines.

There is increasing evidence that carbohydrate-binding proteins (lectins) are widespread in mammals (Kilpatrick, 2002), and especially in cells of the immune system. Several tenths calcium-dependent lectins (C-type lectins; Dodd and Drickamer, 2001; Drickamer, 1997) have been identified each endowed with a different carbohydrate-binding specificity. For example, L-selectin (Foxall et al., 1992; Lasky, 1992) recognizes the 6′-sulphated sialyl-Lewisx epitope. Mannan-binding lectins constitute a family of related calcium-dependent lectins with important implications in the field of human pathology (Kawasaki, 1999; Kilpatrick, 2002). Likewise the calcium-independent lectins comprise the lactose-binding galectins (Cooper, 2002), the I-type lectins (Angata and Brinkman-van der Linden, 2002; Kelm et al., 1994; Nath et al., 1995; Powell and Varki, 1994) and CSL, which is specific for oligomannosidic N-glycans with 6 mannose residues (Zanetta et al., 1987). The MR60-ERGIC-53 mannose-binding lectin (Arar et al., 1995; Fiedler and Simmons, 1994) is related to plant lectins. Besides these molecules involved either in homotypic or heterotypic cell adhesion, or in intracellular traffic of glycoconju-gates, those that are polyvalent (having two carbohydrate-recognition domains (CRD) or organized as oligomers) may have clustering effects on their ligands. This may represent the endogenous mechanism mimicked by specific polyvalent antibodies or plant lectins (Feizi and Childs, 1987). For example, the lectin CSL, which is rapidly over-expressed and externalized after stimulation of human cells with phorbol esters and binds to its surface ligands (including the glycosylated forms of CD3 on T cells and CD24 on B cells) is responsible for the major tyrosine-phosphorylation changes occurring in the early stages of cell activation (Zanetta et al., 1995).

High-resolution Crystal Structures of Erythrina cristagalli Lectin in Complex with Lactose and 2′-α- 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′-α- l-fucosyllactose were determined at 1.6 Å and 1.7 Å 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′-α- l-fucosyllactose, and with two modeled complexes of ECorL with 2′-α- l-fucosyl- N-acetyllactosamine. The molecular models are very similar to the crystal structure of ECL in complex with 2′-α- 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.

The mannose-dependent epitope for neutralizing antibody 2G12 on human immunodeficiency virus type 1 glycoprotein gp120.

We have analyzed the unique epitope for the broadly neutralizing human monoclonal antibody (MAb) 2G12 on the gp120 surface glycoprotein of human immunodeficiency virus type 1 (HIV-1). Sequence analysis, focusing on the conservation of relevant residues across multiple HIV-1 isolates, refined the epitope that was defined previously by substitutional mutagenesis (A. Trkola, M. Purtscher, T. Muster, C. Ballaun, A. Buchacher, N. Sullivan, K. Srinivasan, J. Sodroski, J. P. Moore, and H. Katinger, J. Virol. 70:1100-1108, 1996). In a biochemical study, we digested recombinant gp120 with various glycosidase enzymes of known specificities and showed that the 2G12 epitope is lost when gp120 is treated with mannosidases. Computational analyses were used to position the epitope in the context of the virion-associated envelope glycoprotein complex, to determine the variability of the surrounding surface, and to calculate the surface accessibility of possible glycan- and polypeptide-epitope components. Together, these analyses suggest that the 2G12 epitope is centered on the high-mannose and/or hybrid glycans of residues 295, 332, and 392, with peripheral glycans from 386 and 448 on either flank. The epitope is mannose dependent and composed primarily of carbohydrate, with probably no direct involvement of the gp120 polypeptide surface. It resides on a face orthogonal to the CD4 binding face, on a surface proximal to, but distinct from, that implicated in coreceptor binding. Its conservation amidst an otherwise highly variable gp120 surface suggests a functional role for the 2G12 binding site, perhaps related to the mannose-dependent attachment of HIV-1 to DC-SIGN or related lectins that facilitate virus entry into susceptible target cells.

Lectin Receptor Kinases in Plants

Referee: Dr. Philip Becraft, Zoology and Genetics/Agronomy Depts., 2116 Molecular Building, lowa State University, Ames, IA 50011Forty-two lectin receptor kinase (lecRK)-related sequences and nine related soluble legume lectin sequences were identified in the Arabidopsis thaliana genome. The genes are scattered as a single or gathered copies at different loci throughout the five chromosomes, and four predicted lecRK probably correspond to pseudogenes. Both structural alignments and molecular modeling revealed striking similarities between the lectinlike domain of lecRK, and related A. thaliana soluble lectins and legume lectins. The hydrophobic cavity is extremely conserved, whereas most of the residues forming the monosaccharide-binding site and the bivalent cation-binding site of legume lectins are poorly conserved. LecRK should be unable to bind the simple sugars usually recognized by genuine legume lectins. Molecular modeling of the kinase domain suggests that, except for two apparently inactive receptors, all other lecRK contain a putative functional Ser/Thr kinase catalytic domain. Both the juxtamembrane and C-terminal domains, which are considered important regions for regulating the kinase activity, exhibit a few specific stretches of amino acid residues. Some phylogenetic relationships are inferred from the phylogenetic trees built up from the different lecRK domain sequences. LecRK cluster in three distinct classes (A,B,C), one of them (B) being more closely related to soluble lectins of A. thaliana and legume lectins.

Purification and partial characterization of a lectin from Canavalia grandiflora benth. seeds.

A D-glucose/D-mannose specific lectin from seeds of Canavalia grandiflora (ConGF) was purified by affinity chromatography on Sephadex G-50. By SDS-PAGE ConGF yielded three protein bands with apparent molecular masses of 29-30 kDa (alpha chain), 16-18 kDa (beta fragment) and 12-13 kDa (gamma fragment), like other related lectins from the genus Canavalia (Leguminosae). ConGF strongly agglutinates rabbit erythrocytes, has a high content of ASP and SER, and its N-terminal sequence (30 residues) is highly similar to the sequences of other related lectins from subtribe Diocleinae.