A detailed picture of a protein–carbohydrate hydrogen-bonding network revealed by NMR and MD simulations

Abstract

Cyanovirin-N (CV-N) is a cyanobacterial lectin with antiviral activity towards HIV and several other viruses. Here, we identify mannoside hydroxyl protons that are hydrogen bonded to the protein backbone of the CV-N domain B binding site, using NMR spectroscopy. For the two carbohydrate ligands Manα(1 → 2)ManαOMe and Manα(1 → 2) Manα(1 → 6)ManαOMe five hydroxyl protons, each, are involved in hydrogen-bonding networks. Comparison with previous crystallographic results revealed that four of these hydroxyl protons donate hydrogen bonds to protein backbone carbonyl oxygens in solution and in the crystal. Hydrogen bonds were not detected between the side chains of Glu41 and Arg76 with sugar hydroxyls, as previously proposed for CV-N binding of mannosides. Molecular dynamics simulations of the CV-N/Manα(1 → 2)Manα(1 → 6)ManαOMe complex confirmed the NMR-determined hydrogen-bonding network. Detailed characterization of CV-N/mannoside complexes provides a better understanding of lectin-carbohydrate interactions and opens up to the use of CV-N and similar lectins as antiviral agents.

Designing of a Recombinant Multi-Epitopes Based Vaccine against Enterococcus mundtii Using Bioinformatics and Immunoinformatics Approaches

“…Hydrogen bonds were mainly formed between the electronegative charge particles. Hydrogen bonds are non-covalent forces and are formed between electronegative donors and acceptors [ 59 ]. A visual molecular dynamic (VMD) plugin was utilized for counting and the identification of hydrogen bonding between a vaccine molecule and receptors’ molecules formed in simulation, as shown in Figure 9 C. The threshold value distance value was 3 Å.…”

Glycosidic α-linked mannopyranose disaccharides: an NMR spectroscopy and molecular dynamics simulation study employing additive and Drude polarizable force fields

“…Thus, the present study represents an important step forward in obtaining a more detailed understanding of the conformational properties of mannose saccharides that builds upon four decades of combing experimental methods with molecular modeling. Furthermore, future applications to complex structures or assemblies such as in studies of carbohydrateprotein interactions [126][127][128] may be aided by the improved description of the physical forces dictating the conformational properties obtained with the Drude polarizable force field model thereby representing molecular interactions in a more realistic way.…”

Glycan Shape, Motions, and Interactions Explored by NMR Spectroscopy

“…In contrast, the interactions between the lectin DC-SIGN and 13 C-labeled M9 studied by 1 H, 13 C-HSQC spectroscopy revealed that upon formation of a 1:1 complex between ligand and protein, signals from sugar residues at the core region common to all N-linked oligomannose structures, i.e., in this case α-D-Manp-(1 → 6)β-D-Manp-(1 → 4)-β-D-GlcpNAc-(1 → 4)-D-GlcpNAc, disappeared thereby indicating the binding region and that the binding process takes place on a slower time scale of ∼10 ms. 91 Chemical synthesis using 13 C-uniformly labeled monosaccharides is an alternative to obtain well-defined oligosaccharides and this approach was used to make α-D-Manp-(1 → 2)-α-D-Manp-(1 → 6)-α-D-Manp-OMe (M26). 75 The complex between 15 Nlabeled cyanovirin-N (CV−N) and M26 was studied by 1 H, 13 C-CT-HSQC, 1 H, 13 C-HSQC-NOESY and 1 H, 13 C-HSQC-TOCSY NMR experiments with focus on hydroxyl proton torsion angles of the ligand in the bound conformation, which revealed that specific rotamers of HO3 and HO4 hydroxyl groups in the terminal and central sugar residues were preferred in the CV−N:M26 complex. Enzymatic synthesis was employed for poly-N-acetyllactosamines, which have a repetitive structure of →3)-α-D-Galp-(1 → 4)-β-D-GlcpNAc-(1→ disaccharide entities, with up to hexasaccharides having 13 C-uniformly labeled galactose residues at different positions in the sequence of alternating sugar residues and was facilitated by β1,3-Nacetylglucosaminyltransferase in the presence of UDP-GlcNAc and β1,4-galactosyltransferase in conjunction with UDP-UL-13 C 6 -galactose.…”

“…Glycan–protein interactions are determined by, inter alia, CH−π interactions, , stereoelectronics, and hydrogen-bonding networks . These interactions are usually investigated by more than one experimental technique.…”