Activity–structure correlations in divergent lectin evolution: fine specificity of chicken galectin CG-14 and computational analysis of flexible ligand docking for CG-14 and the closely related CG-16 (original) (raw)
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Glycobiology, 2018
Galectin-10 (Gal-10) which forms Charcot-Leyden crystals in vivo, is crucial to regulating lymph cell function. Here, we solved the crystal structures of Gal-10 and eight variants at resolutions of 1.55-2.00 Å. Structural analysis and size exclusion chromatography demonstrated that Gal-10 dimerizes with a novel global shape that is different from that of other prototype galectins (e.g., Gal-1, -2 and -7). In the Gal-10 dimer, Glu33 from one subunit modifies the carbohydrate-binding site of another, essentially inhibiting disaccharide binding. Nevertheless, glycerol (and possibly other small hydroxylated molecules) can interact with residues at the ligand binding site, with His53 being the most crucial for binding. Alanine substitution of the conserved Trp residue (Trp72) that is crucial to saccharide binding in other galectins, actually leads to enhanced erythrocyte agglutination, suggesting that Trp72 negatively regulates Gal-10 ligand binding. Overall, our crystallographic and bio...
Journal of Molecular Modeling, 1997
Galectins (Galactose binding lectins) from bacteria, plants and animals have been shown to possess tyrosine or tryptophan residues that form hydrophobic contacts with their ligands in the binding sites. At the present time, the X-ray structures of only two galectins from human and bovine tissues are known. In the present study we applied X-ray data of bovine heart galectin-1 as a template for homology modelling of a number of galectins from mammalian and avian tissues. The conservation of one tryptophan and at least one histidine in binding pocket can be observed from the comparison of the model structures. We also show that it is possible to obtain information of the architecture of the binding pocket of several galectins in solution using CIDNP (Chemically Induced Dynamic Nuclear Polarisation) techniques. The CIDNP approach offers a possibility to analyse these lectins in solution thereby providing supplementary information to the available X-ray data. All studied galectins show comparable alterations when they are recorded by CIDNPtechnique in the absence and in the presence of their specific carbohydrate ligands.
Glycobiology, 2000
of the carbohydrate recognition domain (CRD, amino acid residues 114-245) of hamster galectin-3 has been extended to include N-terminal domain amino acid residues 91-113 containing one of the nine proline-rich motifs present in full-length hamster galectin-3. The modeling predicts two configurations of the N-terminal tail: in one the tail turns toward the first (SI) and last (S12) β-strands of the CRD and lies at the apolar dimer interface observed for galectins-1 and-2. In the second folding arrangement the N-terminal tail lies across the carbohydrate-binding pocket of the CRD where it could participate in sugar-binding: in particular tyrosine 102 and adjacent residues may interact with the partly solvent exposed nonreducing N-acetylgalactosamine and fucose substituents of the A-blood group structure GalNAcα1,3 [Fucα1,2]Galβ1,4GlcNAc-R. Binding studies using surface plasmon resonance of a recombinant fragment ∆1-93 protein containing residues 94-245 of hamster galectin-3 and a collagenase-derived fragment ∆1-103 containing residues 104-245, as well as alanine mutagenesis of residues 101-105 in ∆1-93 protein, support the prediction that Tyr102 and adjacent residues make significant contributions to oligosaccharide binding.
Combining Crystallography and Hydrogen-Deuterium Exchange to Study Galectin-Ligand Complexes
Chemistry: A European Journal, 2015
The physiological significance arising from translating information stored in glycans into cellular effects explains the interest in structurally defining lectin-carbohydrate recognition. The relatively smalls et of adhesion/ growth-regulatory galectins in chicken makes this system attractive to study the origins of specificity and divergence. Cell binding by using glycosylation mutantsr eveals binding of the N-terminal domain of chickeng alectin-8 (CG-8N) to a-2,3-sialylated and galactose-terminated glycanc hains. Cocrystals with lactose and its 3'-sialylated derivatived isclose Arg58 as ak ey contact for the carboxylic acida nd differences in loop lengths to the three homodimericc hickeng alectins. Monitoring hydrogen-deuterium exchangeb ym ass spectrometry revealed an effective reduction of deuteration after ligand binding within the contact area.I na ddition, evidence for changes in solvent accessibility of amide protons beyondt his site was obtained. Their detection, which highlights the sensorc apacity of this technique, encourages systematicstudies on galectins and beyond.
Glycobiology, 2016
Endogenous lectins can control critical biological responses, including cell communication, signaling, angiogenesis and immunity by decoding glycan-containing information on a variety of cellular receptors and the extracellular matrix. Galectin-1 (Gal-1), a prototype member of the galectin family, displays only one carbohydrate recognition domain and occurs in a subtle homodimerization equilibrium at physiologic concentrations. Such equilibrium critically governs the function of this lectin signaling by allowing tunable interactions with a preferential set of glycosylated receptors. Here, we used a combination of experimental and computational approaches to analyze the kinetics and mechanisms connecting Gal-1 ligand unbinding and dimer dissociation processes. Kinetic constants of both processes were found to differ in an order of magnitude. By means of steered molecular dynamics simulation, the ligand unbinding process was followed in combination with water occupancy changes. By det...
Glycobiology, 2006
Cell surface glycans are functional docking sites for tissue lectins such as the members of the galectin family. This interaction triggers a wide variety of responses; hence, there is a keen interest in defining its structural features. Toward this aim, we have used enzyme-linked lectinosorbent and inhibition assays with the proto-type rat galectin-5 and panels of free saccharides and glycoconjugates. Among 45 natural glycans tested for lectin binding, galectin-5 reacted best with glycoproteins (gps) presenting a high density of Galβ1-3/4GlcNAc (I/II) and multi-antennary N-glycans with II termini. Their reactivities, on a nanogram basis, were up to 4.3×10 2 , 3.2×10 2 , 2.5×10 2 and 1.7×10 4 times higher than monomeric Galβ1-3/4GlcNAc (I/II), triantennary-II (Tri-II) and Gal, respectively. Galectin-5 also bound well to several blood-group type B (Galα1-3Gal)-and A (GalNAcα1-3Gal)-containing gps. It reacted weakly or not at all with tumor-associated Tn (GalNAcα1-Ser/Thr) and sialylated gps.
PLoS ONE, 2013
Human Galectin-8 (Gal-8) is a member of the galectin family which shares an affinity for b-galactosides. The tandem-repeat Gal-8 consists of a N-and a C-terminal carbohydrate recognition domain (N-and C-CRD) joined by a linker peptide of various length. Despite their structural similarity both CRDs recognize different oligosaccharides. While the molecular requirements of the N-CRD for high binding affinity to sulfated and sialylated glycans have recently been elucidated by crystallographic studies of complexes with several oligosaccharides, the binding specificities of the C-CRD for a different set of oligosaccharides, as derived from experimental data, has only been explained in terms of the three-dimensional structure for the complex C-CRD with lactose. In this study we performed molecular dynamics (MD) simulations using the recently released crystal structure of the Gal-8C-CRD to analyse the three-dimensional conditions for its specific binding to a variety of oligosaccharides as previously defined by glycan-microarray analysis. The terminal b-galactose of disaccharides (LacNAc, lacto-N-biose and lactose) and the internal b-galactose moiety of blood group antigens A and B (BGA, BGB) as well as of longer linear oligosaccharide chains (di-LacNAc and lacto-N-neotetraose) are interacting favorably with conserved amino acids (H53, R57, N66, W73, E76). Lacto-N-neotetraose and di-LacNAc as well as BGA and BGB are well accommodated. BGA and BGB showed higher affinity than LacNAc and lactose due to generally stronger hydrogen bond interactions and water mediated hydrogen bonds with a1-2 fucose respectively. Our results derived from molecular dynamics simulations are able to explain the glycan binding specificities of the Gal-8C-CRD in comparison to those of the Gal-8N-CRD.
Influence of protein (human galectin-3) design on aspects of lectin activity
Histochemistry and Cell Biology, 2020
The concept of biomedical significance of the functional pairing between tissue lectins and their glycoconjugate counterreceptors has reached the mainstream of research on the flow of biological information. A major challenge now is to identify the principles of structure–activity relationships that underlie specificity of recognition and the ensuing post-binding processes. Toward this end, we focus on a distinct feature on the side of the lectin, i.e. its architecture to present the carbohydrate recognition domain (CRD). Working with a multifunctional human lectin, i.e. galectin-3, as model, its CRD is used in protein engineering to build variants with different modular assembly. Hereby, it becomes possible to compare activity features of the natural design, i.e. CRD attached to an N-terminal tail, with those of homo- and heterodimers and the tail-free protein. Thermodynamics of binding disaccharides proved full activity of all proteins at very similar affinity. The following glyca...
Monovalent Interactions of Galectin-1
Biochemistry, 2010
Galectin-1, a β-galactoside binding lectin involved in immunoregulation and cancer, binds natural and many synthetic multivalent glycoconjugates with an apparent glycoside cluster effect, that is, affinity above and beyond what would be expected from the concentration of the determinant sugar. Here we have analyzed the mechanism of such cluster effects in solution at physiological concentration using a fluorescence anisotropy assay with a novel fluorescent high-affinity galectin-1 binding probe. The interaction of native dimeric and monomeric mutants of rat and human galectin-1 with mono-and divalent small molecules, fetuin, asialofetuin, and human serum glycoproteins was analyzed. Surprisingly, high-affinity binding did not depend much on the dimeric state of galectin-1 and thus is due mainly to monomeric interactions of a single carbohydrate recognition domain. The mechanism for this is unknown, but one possibility includes additional interactions that high-affinity ligands make with an extended binding site on the carbohydrate recognition domain. It follows that such weak additional interactions must be important for the biological function of galectin-1 and also for the design of galectin-1 inhibitors.