Theoretical investigation on the glycan-binding specificity of Agrocybe cylindracea galectin using molecular modeling and molecular dynamics simulation studies.

Galectins are ß-galactoside binding proteins which have the ability to serve as potent antitumor, cancer biomarker, and induce tumor cell apoptosis. Agrocybe cylindracea galectin (ACG) is a fungal galectin which specifically recognizes α(2,3)-linked sialyllactose at the cell surface that plays extensive roles in the biological recognition processes. To investigate the change in glycan-binding specificity upon mutations, single point and double point site-directed in silico mutations are performed at the binding pocket of ACG. Molecular dynamics (MD) simulation studies are carried out for the wild-type (ACG) and single point (ACG1) and double point (ACG2) mutated ACGs to investigate the dynamics of substituted mutants and their interactions with the receptor sialyllactose. Plausible binding modes are proposed for galectin-sialylglycan complexes based on the analysis of hydrogen bonding interactions, total pair-wise interaction energy between the interacting binding site residues and sialyllactose and binding free energy of the complexes using molecular mechanics-Poisson-Boltzmann surface area. Our result shows that high contribution to the binding in different modes is due to the direct and water-mediated hydrogen bonds. The binding specificity of double point mutant Y59R/N140Q of ACG2 is found to be high, and it has 26 direct and water-mediated hydrogen bonds with a relatively low-binding free energy of -47.52 ± 5.2 kcal/mol. We also observe that the substituted mutant Arg59 is crucial for glycan-binding and for the preference of α(2,3)-linked sialyllactose at the binding pocket of ACG2 galectin. When compared with the wild-type and single point mutant, the double point mutant exhibits enhanced affinity towards α(2,3)-linked sialyllactose, which can be effectively used as a model for biological cell marker in cancer therapeutics.

Insights into the binding specificity of wild type and mutated wheat germ agglutinin towards Neu5Acα(2-3)Gal: a study by in silico mutations and molecular dynamics simulations.

Wheat germ agglutinin (WGA) is a plant lectin, which specifically recognizes the sugars NeuNAc and GlcNAc. Mutated WGA with enhanced binding specificity can be used as biomarkers for cancer. In silico mutations are performed at the active site of WGA to enhance the binding specificity towards sialylglycans, and molecular dynamics simulations of 20 ns are carried out for wild type and mutated WGAs (WGA1, WGA2, and WGA3) in complex with sialylgalactose to examine the change in binding specificity. MD simulations reveal the change in binding specificity of wild type and mutated WGAs towards sialylgalactose and bound conformational flexibility of sialylgalactose. The mutated polar amino acid residues Asn114 (S114N), Lys118 (G118K), and Arg118 (G118R) make direct and water mediated hydrogen bonds and hydrophobic interactions with sialylgalactose. An analysis of possible hydrogen bonds, hydrophobic interactions, total pair wise interaction energy between active site residues and sialylgalactose and MM-PBSA free energy calculation reveals the plausible binding modes and the role of water in stabilizing different binding modes. An interesting observation is that the binding specificity of mutated WGAs (cyborg lectin) towards sialylgalactose is found to be higher in double point mutation (WGA3). One of the substituted residues Arg118 plays a crucial role in sugar binding. Based on the interactions and energy calculations, it is concluded that the order of binding specificity of WGAs towards sialylgalactose is WGA3 > WGA1 > WGA2 > WGA. On comparing with the wild type, double point mutated WGA (WGA3) exhibits increased specificity towards sialylgalactose, and thus, it can be effectively used in targeted drug delivery and as biological cell marker in cancer therapeutics.

Theoretical investigation on the binding specificity of sialyldisaccharides with hemagglutinins of influenza A virus by molecular dynamics simulations.

Recognition of cell-surface sialyldisaccharides by influenza A hemagglutinin (HA) triggers the infection process of influenza. The changes in glycosidic torsional linkage and the receptor conformations may alter the binding specificity of HAs to the sialylglycans. In this study, 10-ns molecular dynamics simulations were carried out to examine the structural and dynamic behavior of the HAs bound with sialyldisaccharides Neu5Acα(2-3)Gal (N23G) and Neu5Acα(2-6)Gal (N26G). The analysis of the glycosidic torsional angles and the pair interaction energy between the receptor and the interacting residues of the binding site reveal that N23G has two binding modes for H1 and H5 and a single binding mode for H3 and H9. For N26G, H1 and H3 has two binding modes, and H5 and H9 has a single binding mode. The direct and water-mediated hydrogen bonding interactions between the receptors and HAs play dominant roles in the structural stabilization of the complexes. It is concluded from pair interaction energy and Molecular Mechanic-Poisson-Boltzmann Surface Area calculations that N26G is a better receptor for H1 when compared with N23G. N23G is a better receptor for H5 when compared with N26G. However, H3 and H9 can recognize N23G and N26G in equal binding specificity due to the marginal energy difference (≈2.5 kcal/mol). The order of binding specificity of N23G is H3 > H5 > H9 > H1 and N26G is H1 > H3 > H5 > H9, respectively. The proposed conformational models will be helpful in designing inhibitors for influenza virus.

Sialyldisaccharide conformations: a molecular dynamics perspective.

Sialyldisaccharides are significant terminal components of glycoconjugates and their negative charge and conformation are extensively utilized in molecular recognition processes. The conformation and flexibility of four biologically important sialyldisaccharides [Neu5Acα(2-3)Gal, Neu5Acα(2-6)Gal, Neu5Acα(2-8)Neu5Ac and Neu5Acα(2-9)Neu5Ac] are studied using Molecular Dynamics simulations of 20 ns duration to deduce the conformational preferences of the sialyldisaccharides and the interactions which stabilize the conformations. This study clearly describes the possible conformational models of sialyldisaccharides deduced from 20 ns Molecular Dynamics simulations and our results confirm the role of water in the structural stabilization of sialyldisaccharides. An extensive analysis on the sialyldisaccharide structures available in PDB also confirms the conformational regions found by experiments are detected in MD simulations of 20 ns duration. The three dimensional structural coordinates for all the MD derived sialyldisaccharide conformations are deposited in the 3DSDSCAR database and these conformational models will be useful for glycobiologists and biotechnologists to understand the biological functions of sialic acid containing glycoconjugates.

Molecular dynamics simulation and quantum mechanical calculations on α-D-N-acetylneuraminic acid.

N-Acetylneuraminic acid is a sugar molecule of biological significance due to its pivotal role in molecular recognition processes. The three dimensional structure and conformation of α-Neu5Ac in biological environments can be clearly observed by molecular dynamics (MD) simulation and quantum mechanical (QM) calculations. A 10ns MD simulation on α-Neu5Ac yields two conformational models which are stabilized by water mediated hydrogen bond between O-8/O-9 hydroxyl oxygen and carbonyl of carboxylate group. The average life time of the conformers and the residual time of water which mediates the hydrogen bonding interactions are computed. Based on the amphiprotic nature of water, water mediation of each conformer is divided into two different modes, one donor-one acceptor mode and two donor modes. According to the analysis of simulation trajectories, the preferred mode of water mediation for conformers is the one donor-one acceptor mode. The energy and geometry of the MD derived conformational models of α-Neu5Ac are optimized using HF/6-31G(∗) basis set of Gaussian03. QM calculations also resulted that α-Neu5Ac is preferentially stabilized by water mediated hydrogen bonding between O-8 hydroxyl and the carboxylate group where the mediation is one donor-one acceptor type. The optimized geometry of α-Neu5Ac which is in good agreement with the crystal structure of α-D-N-acetyl-1-O-methylneuraminic acid methyl ester is deposited in the public domain database 3DSDSCAR (http://3dsdscar.org). This optimized structure can be used by biotechnologists, biophysicists and glycobiologists for modelling the sialylglycans and also to design drugs using sialic acid analog inhibitors.

3DSDSCAR--a three dimensional structural database for sialic acid-containing carbohydrates through molecular dynamics simulation.

The inherent flexibility and lack of strong intramolecular interactions of oligosaccharides demand the use of theoretical methods for their structural elucidation. In spite of the developments of theoretical methods, not much research on glycoinformatics is done so far when compared to bioinformatics research on proteins and nucleic acids. We have developed three dimensional structural database for a sialic acid-containing carbohydrates (3DSDSCAR). This is an open-access database that provides 3D structural models of a given sialic acid-containing carbohydrate. At present, 3DSDSCAR contains 60 conformational models, belonging to 14 different sialic acid-containing carbohydrates, deduced through 10 ns molecular dynamics (MD) simulations. The database is available at the URL: http://www.3dsdscar.org.