Conserved Carbohydrate Recognition Domain Research Articles

Galectins in cell growth and apoptosis.

Fourteen members of the galectin family, proteins with conserved carbohydrate-recognition domains that bind beta-galactoside, have been cloned and more are expected to be discovered in the near future. Many aspects of galectin biology have been thoroughly explored, and functional studies have implicated these proteins in cell growth, differentiation and apoptosis, in addition to cell adhesion, chemoattraction and cell migration. In some cases a galectin can either promote or suppress cell growth, depending on the cell types and doses used. Galectin-3 is the only member known so far to inhibit apoptosis, while galectin-1, -7 and -9 promote this cellular process. Galectins can act either extracellularly or intracellularly to exert effects on cell growth and apoptosis.

Biological activities of ecalectin: a novel eosinophil-activating factor.

Ecalectin, produced by Ag-stimulated T lymphocytes, is a potent eosinophil-specific chemoattractant in vitro as well as in vivo and thus is implicated in allergic responses. Ecalectin differs structurally from other known eosinophil chemoattractants (ECAs); ecalectin belongs to the galectin family defined by their affinity for beta-galactosides and by their conserved carbohydrate recognition domains. These characteristic features suggest that ecalectin has unique activities associated with allergic inflammation besides ECA activity. Conversely, ecalectin may mediate ECA activity by binding to a receptor of a known ECA via affinity for the beta-galactosides present on this receptor. In this study, we have tested whether ecalectin mediates ECA activity by binding to a receptor of a known ECA, and we have assessed its effects on eosinophils. Ecalectin did not mediate ECA activity by binding to the IL-5R or to CCR3. Also, the ECA activity of ecalectin was mainly chemokinetic. In addition, ecalectin induced concentration-dependent eosinophil aggregation, a marker for eosinophil activation. Ecalectin induced concentration-dependent superoxide production from eosinophils but did not induce degranulation; usually these two events are coupled in eosinophil activation. Moreover, ecalectin directly prolonged eosinophil survival in vitro and did not trigger eosinophils to secrete cytokines that prolong eosinophil survival. These results demonstrate that ecalectin has several unique effects on eosinophils. Therefore, we conclude that ecalectin is a novel eosinophil-activating factor. Presumably, these effects allow ecalectin to play a distinctive role in allergic inflammation.

Homology modelling and molecular dynamics studies of human placental tissue protein 13 (galectin-13).

The primary structure of the newly sequence analysed placental tissue protein 13 (PP13) was highly homologous to several members of the beta-galactoside-binding S-type lectin (galectin) family. By homology modelling, the three-dimensional structure of PP13 was built based on high-resolution crystal structures of homologues and also their characteristic 'jellyroll' fold was found in the case of PP13. Our model has been deposited in the Brookhaven Protein Data Bank. By multiple sequence alignment and structure-based secondary structure prediction, we underlined the structural similarity of PP13 with its homologues. The secondary structure of PP13 was identical with 'proto-type' galectins consisting of a five- and a six-stranded beta-sheet, joined by two alpha-helices, and galectins' highly conserved carbohydrate-recognition domain (CRD) was also present in PP13. Of the eight consensus residues in the CRD, four identical and three conservatively substituted were shared by PP13. By docking simulations PP13 possessed sugar-binding activity with highest affinity to N-acetyllactosamine and lactose typical of most galectins. All ligands were docked into the putative CRD of PP13. Based on several lines of evidence discussed in this paper demonstrating that PP13 is a novel galectin, PP13 was also designated galectin-13. These computational results provide some new insights into the possible role and importance of PP13 in various processes of the human body and can be of help in the initial steps of further functional research.

Cell Cycle Regulation by Galectin-12, a New Member of the Galectin Superfamily

Galectins are a family of beta-galactoside-binding animal lectins with conserved carbohydrate recognition domains (CRDs). Here we report the identification and characterization of a new galectin, galectin-12, which contains two domains that are homologous to the galectin CRD. The N-terminal domain contains all of the sequence elements predicted to form the two beta-sheets found in other galectins, as well as conserved carbohydrate-interacting residues. The C-terminal domain shows considerable divergence from the consensus sequence, and many of these conserved residues are not present. Nevertheless, the protein has lactose binding activity, most likely due to the contribution of the N-terminal domain. The mRNA for galectin-12 contains features coding for proteins with growth-regulatory functions. These include start codons in a context that are suboptimal for translation initiation and AU-rich motifs in the 3'-untranslated region, which are known to confer instability to mRNA. Galectin-12 mRNA is sparingly expressed or undetectable in many tissues and cell lines tested, but it is up-regulated in cells synchronized at the G(1) phase or the G(1)/S boundary of the cell cycle. Ectopic expression of galectin-12 in cancer cells causes cell cycle arrest at the G(1) phase and cell growth suppression. We conclude that galectin-12 is a novel regulator of cellular homeostasis.

Requirement of Divalent Galactoside-binding Activity of Ecalectin/Galectin-9 for Eosinophil Chemoattraction

We have previously isolated and cloned a novel eosinophil chemoattractant (ECA) from a human T-cell-derived expression library. This ECA, termed ecalectin, is a variant of human galectin-9, a member of a beta-galactoside binding animal lectin family, which contains two conserved carbohydrate recognition domains (CRDs). In the present study, we addressed whether carbohydrate binding activity is required for the ECA activity of ecalectin and whether both CRDs are essential for this activity. Recombinant full-length wild-type ecalectin (ecalectin-WT) and N-terminal and C-terminal CRD (ecalectin-NT and -CT, respectively) were generated. All of these recombinant proteins exhibited affinity for lactose, a property shared by galectins, but ecalectin-WT exhibited substantially higher hemagglutination activities than ecalectin-NT and -CT. Furthermore, ecalectin-WT showed over 100-fold higher ECA activity than ecalectin-NT and -CT; combination of recombinant domain fragments did not reconstitute the ECA and hemagglutination activities of the full-length protein. ECA activity of ecalectin-WT was inhibited by lactose in a dose-dependent manner. Site-directed mutation of positions Arg(65) of ecalectin-NT and Arg(239) of ecalectin-CT to an aspartic acid residue resulted in the loss of both lactose-binding and ECA activities. We conclude that divalent galactoside-binding activity is required for eosinophil chemoattraction by ecalectin.

Ca2+-dependent structural changes in C-type mannose-binding proteins.

C-type animal lectins are a diverse family of proteins which mediate cell-surface carbohydrate-recognition events through a conserved carbohydrate-recognition domain (CRD). Most members of this family possess a carbohydrate-binding activity that depends strictly on the binding of Ca2+ at two sites, designated 1 and 2, in the CRD. The structural transitions associated with Ca2+ binding in C-type lectins have been investigated by determining high-resolution crystal structures of rat serum mannose-binding protein (MBP) bound to one Ho3+ in place of Ca2+, and the apo form of rat liver MBP. The removal of Ca2+ does not affect the core structure of the CRD, but dramatic conformational changes occur in the loops. The most significant structural change in the absence of Ca2+ is the isomerization of a cis-peptide bond preceding a conserved proline residue in Ca2+ site 2. This bond adopts the cis conformation in all Ca2+-bound structures, whereas both cis and trans conformations are observed in the absence of Ca2+. The pattern of structural changes in the three loops that interact with Ca2+ is dictated in large part by the conformation of the prolyl peptide bond. The highly conserved nature of Ca2+ site 2 suggests that the transitions observed in MBPs are general features of Ca2+ binding in C-type lectins.