467 research outputs found
Structural basis for the carbohydrate specificities of artocarpin: variation in the length of a loop as a strategy for generating ligand specificity
Artocarpin, a tetrameric lectin of molecular mass 65 kDa, is one of the two lectins extracted from the seeds of jackfruit. The structures of the complexes of artocarpin with mannotriose and mannopentose reported here, together with the structures of artocarpin and its complex with Me-α-mannose reported earlier, show that the lectin possesses a deep-seated binding site formed by three loops. The binding site can be considered as composed of two subsites; the primary site and the secondary site. Interactions at the primary site composed of two of the loops involve mainly hydrogen bonds, while those at the secondary site comprising the third loop are primarily van der Waals in nature. Mannotriose in its complex with the lectin interacts through all the three mannopyranosyl residues; mannopentose interacts with the protein using at least three of the five mannose residues. The complexes provide a structural explanation for the carbohydrate specificities of artocarpin. A detailed comparison with the sugar complexes of heltuba, the only other mannose-specific jacalin-like lectin with known three-dimensional structure in sugar-bound form, establishes the role of the sugar-binding loop constituting the secondary site, in conferring different specificities at the oligosaccharide level. This loop is four residues longer in artocarpin than in heltuba, providing an instance where variation in loop length is used as a strategy for generating carbohydrate specificity
A23187-Channel behaviour: fluorescence study
Pyranine entrapped soylipid liposomes have been used as a model system to study the proton transport across membrane in the presence of A23187, a carboxylic ionophore specific for electroneutral exchange of divalent cations. An apparent rate constant (kapp) for transport of protons has been determined from the rate of change of fluorescence intensity of pyranine by stopped flow rapid kinetics in the presence of proton gradient The variation of kapp has been studied as a function of ionophore concentration and the results have been compared with gramicidin-a well known channel former under the similar experimental conditions. The rates thus obtained showed that A23187 is not only a simple carrier but also shows channel behaviour at high concentration of ionophore
Fluorescence polarization as a tool to study lectin-sugar interaction
The binding of Ricinus communis agglutinin and Abrus agglutinin to 4-methylumbelliferyl β-D-galactopyranoside was studied by equilibrium dialysis, fluo-rescence quenching and fluorescence polarization. The number of binding sites and the association constant value obtained by fluorescence polarization for both Ricinus communis agglutinin and Abrus agglutinin are in close agreement with those obtained by the other methods. This indicates the potential of ligand-fluorescence polarization measurements in the investigation of lectin-sugar interactions
Carbohydrate specificity and salt-bridge mediated conformational change in acidic winged bean agglutinin
Structures of two crystal forms of the dimeric acidic winged bean agglutinin (WBAII) complexed with methyl-α-D-galactose have been determined at 3.0 Å and 3.3 Å resolution. The subunit structure and dimerisation of the lectin are similar to those of the basic lectin from winged bean (WBAI) and the lectin from Erythrina corallodendron (EcorL). The conformation of a loop and its orientation with respect to the rest of the molecule in WBAII are, however, different from those in all the other legume lectins of known structure. This difference appears to have been caused by the formation of two strategically placed salt bridges in the former. Modelling based on the crystal structures provides a rationale for the specificity of the lectin, which is very different from that of WBAI, for the H-antigenic determinant responsible for O blood group reactivity. It also leads to a qualitative explanation for the thermodynamic data on sugar-binding to the lectin, with special emphasis on the role of a tyrosyl residue in the variable loop in the sugar-binding region in generating the carbohydrate specificity of WBAII
Re-refinement using reprocessed data to improve the quality of the structure: a case study involving garlic lectin
Structural basis of the carbohydrate specificities of Jacalin: an X-ray and modeling study
The structures of the complexes of tetrameric jacalin with Gal, Me-α-GalNAc, Me-α-T-antigen, GalNAcβ1-3Gal-α-O-Me and Galα1-6Glc (mellibiose) show that the sugar-binding site of jacalin has three components: the primary site, secondary site A, and secondary site B. In these structures and in the two structures reported earlier, Gal or GalNAc occupy the primary site with the anomeric carbon pointing towards secondary site A. The α-substituents, when present, interact, primarily hydrophobically, with secondary site A which has variable geometry. O-H···π and C-H···π hydrogen bonds involving this site also exist. On the other hand, β-substitution leads to severe steric clashes. Therefore, in complexes involving β-linked disaccharides, the reducing sugar binds at the primary site with the non-reducing end located at secondary site B. The interactions at secondary site B are primarily through water bridges. Thus, the nature of the linkage determines the mode of the association of the sugar with jacalin. The interactions observed in the crystal structures and modeling based on them provide a satisfactory qualitative explanation of the available thermodynamic data on jacalin-carbohydrate interactions. They also lead to fresh insights into the nature of the binding of glycoproteins by jacalin
Plasmepsin inhibitors: design, synthesis, inhibitory studies and crystal structure analysis
Plasmepsin group of enzymes are key enzymes in the life cycle of malarial parasites. As inhibition of plasmepsins leads to the parasite's death, these enzymes can be utilized as potential drug targets. Although many drugs are available, it has been observed that Plasmodium falciparum, the species that causes most of the malarial infections and subsequent death, has developed resistance against most of the drugs. Based on the cleavage sites of hemglobin, the substrate for plasmepsins, we have designed two compounds (p-nitrobenzoyl-leucine-β -alanine and p-nitrobenzoyl-leucine-isonipecotic acid), synthesized them, solved their crystal structures and studied their inhibitory effect using experimental and theoretical (docking) methods. In this paper, we discuss the synthesis, crystal structures and inhibitory nature of these two compounds which have a potential to inhibit plasmepsins
Multivalency in lectins-a crystallographic,modelling and light-scattering study involving peanut lectin and a bivalent ligand
Multivalency is believed to be important in the activity of lectins, although definitive structural studies on it have been few and far between. We have now studied the complexation of tetravalent peanut lectin with a synthetic compound containing two terminal lactose moieties, using a combination of crystallography, dynamic light scattering and modelling. Light scattering indicates the formation of an apparent dimeric species and also larger aggregates of the tetrameric lectin in the presence of the bivalent ligand. Crystals of presumably crosslinked lectin molecules could be obtained. They diffract poorly, but X-ray data from them are good enough to define the positions of the lectin molecules. Extensive modelling on possible crosslinking modes of protein molecules by the ligand indicated that systematic crosslinking could lead to crystalline arrays. The studies also provided a rationale for crosslinking in the observed crystal structure. The results obtained provide further insights into the general problem of multivalency in lectins. They indicate that crosslinking involving multivalent lectins and multivalent carbohydrates could lead to an ensemble of a finite number of distinct periodic arrays rather than a unique array
Crystallographic identification of an ordered C-terminal domain and a second nucleotide-binding site in RecA: new insights into allostery
RecA protein is a crucial and central component of the homologous recombination and DNA repair machinery. Despite numerous studies on the protein, several issues concerning its action, including the allosteric regulation mechanism have remained unclear. Here we report, for the first time, a crystal structure of a complex of Mycobacterium smegmatis RecA (MsRecA) with dATP, which exhibits a fully ordered C-terminal domain, with a second dATP molecule bound to it. ATP binding is an essential step for all activities of RecA, since it triggers the formation of active nucleoprotein filaments. In the crystal filament, dATP at the first site communicates with a dATP of the second site of an adjacent subunit, through conserved residues, suggesting a new route for allosteric regulation. In addition, subtle but definite changes observed in the orientation of the nucleotide at the first site and in the positions of the segment preceding loop L2 as well as in the segment 102–105 situated between the 2 nt, all appear to be concerted and suggestive of a biological role for the second bound nucleotide
Structural plasticity of peanut lectin: an X-ray analysis involving variation in pH, ligand binding and crystal structure
Until recently, it has only been possible to grow crystals of peanut lectin when complexed with sugar ligands. It is now shown that it is possible to grow peanut lectin crystals at acidic pH in the presence of oligopeptides corresponding to a loop in the lectin molecule. Crystals have also been prepared in the presence of these peptides as well as lactose. Low-pH crystal forms of the lectin-lactose complex similar to those obtained at neutral pH have also been grown. Thus, crystals of peanut lectin grown under different environmental conditions, at two pH values with and without sugar bound to the lectin, are now available. They have been used to explore the plasticity and hydration of the molecule. A detailed comparison between different structures shows that the lectin molecule is sturdy and that the effect of changes in pH, ligand binding and environment on it is small. The region involving the curved front β-sheet and the loops around the second hydrophobic core is comparatively rigid. The back β-sheet involved in quaternary association, which exhibits considerable variability, is substantially flexible, as is the sugar-binding region. The numbers of invariant water molecules in the hydration shell are small and they are mainly involved in metal coordination or in stabilizing unusual structural features. Small consistent movements occur in the combining site upon sugar binding, although the site is essentially preformed
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