Binding of different monosaccharides by lectin PA-IIL from Pseudomonas aeruginosa: Thermodynamics data correlated with X-ray structures (original) (raw)
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Journal of Biological Chemistry, 2010
The calcium-dependent lectin I from Pseudomonas aeruginosa (PA-IL) binds specifically to oligosaccharides presenting an -galactose residue at their non-reducing end, such as the disaccharides Gal1-2GalOMe, Gal1-3GalOMe and Gal1-4GalOMe. This provides a unique model for studying the effect of the glycosidic linkage of the ligands on structure and thermodynamics of the complexes by means of experimental and theoretical tools. The structural features of PA-IL in complex with the three disaccharides were established by docking and molecular dynamics (MD) simulations and compared to those observed in available crystal structures, including PA-IL/Gal1-2GalOMe complex, that was solved at 2.4 Å resolution and reported herein. The role of a structural bridge water molecule in the binding site of -PAIL was also elucidated through MD simulations and molecular mechanics Poisson-Bolzmann surface area MM-PBSA approach. This water molecule establishes three very stable hydrogen bonds with O6 of non-reducing galactose, oxygen from Pro51 main chain and nitrogen from Gln53 main chain of the lectin binding site. Binding free energies for PA-IL in complex with the three disaccharides were investigated and the results were compared with the experimental data determined by titration microcalorimetry. When the bridge water molecule was included in the MM-PBSA calculations, the simulations predicted the correct binding affinity trends with the 1-2 linked disaccharide presenting three times stronger affinity ligand than the two other ones. These results highlight the role of the water molecule in the binding site of PA-IL and indicate that it should be taken into account when designing glycoderivatives active against P. aeruginosa adhesion.
Biochemical Journal, 2005
One of the mechanisms contributing to the protection by breastfeeding of the newborn against enteric diseases is related to the ability of human milk oligosaccharides to prevent the attachment of pathogenic bacteria to the duodenual epithelium. Indeed, a variety of fucosylated oligosaccharides, specific to human milk, form part of the innate immune system. In the present study, we demonstrate the specific blocking of PA-IIL, a fucose-binding lectin of the human pathogen Pseudomonas aeruginosa, by milk oligosaccharides. Two fucosylated epitopes, Lewis a and 3-fucosyl-lactose (Lewis x glucose analogue) bind to the lectin with dissociation constants of 2.2 × 10 −7 M and 3.6 × 10 −7 M respectively. Thermodynamic studies indicate that these interactions are dominated by enthalpy. The entropy contribution is slightly favourable when binding to fucose and to the highest-affinity ligand, Lewis a. The high-resolution X-ray structures of two complexes of PA-IIL with milk oligosaccharides allow the precise determination of the conformation of a trisaccharide and a pentasaccharide. The different types of interaction between the oligosaccharides and the protein involve not only hydrogen bonding, but also calcium-and waterbridged contacts, allowing a rationalization of the thermodynamic data. This study provides important structural information about compounds that could be of general application in new therapeutic strategies against bacterial infections.
Structural basis of calcium and galactose recognition by the lectin PA-IL ofPseudomonas aeruginosa
FEBS Letters, 2003
The structure of the tetrameric Pseudomonas aeruginosa lectin I (PA-IL) in complex with galactose and calcium was determined at 1.6 A î resolution, and the native protein was solved at 2.4 A î resolution. Each monomer adopts a L L-sandwich fold with ligand binding site at the apex. All galactose hydroxyl groups, except O1, are involved in a hydrogen bond network with the protein and O3 and O4 also participate in the coordination of the calcium ion. The stereochemistry of calcium galactose binding is reminiscent of that observed in some animal C-type lectins. The structure of the complex provides a framework for future design of anti-bacterial compounds.
Journal of Molecular Biology, 2008
The opportunistic pathogen Pseudomonas aeruginosa contains several carbohydrate-binding proteins, among which is the P. aeruginosa lectin I (PA-IL), which displays affinity for α-galactosylated glycans. Glycan arrays were screened and demonstrated stronger binding of PA-IL toward αGal1-4βGal-terminating structures and weaker binding to αGal1-3βGal ones in order to determine which human glycoconjugates could play a role in the carbohydrate-mediated adhesion of the bacteria. This was confirmed in vivo by testing the binding of the lectin to Burkitt lymphoma cells that present large amounts of globotriaosylceramide antigen Gb3/CD77/P k . Trisaccharide moieties of Gb3 (αGal1-4βGal1-4Glc) and isoglobotriaosylceramide (αGal1-3βGal1-4Glc) were tested by titration microcalorimetry, and both displayed similar affinity to PA-IL in solution. The crystal structure of PA-IL complexed to αGal1-3βGal1-4Glc trisaccharide has been solved at 1.9-Ǻ resolution and revealed how the second galactose residue makes specific contacts with the protein surface. Molecular modeling studies were performed in order to compare the binding mode of PA-IL toward αGal1-3Gal with that toward αGal1-4Gal. Docking studies demonstrated that αGal1-4Gal creates another network of contacts for achieving a very similar affinity, and 10-ns molecular dynamics in explicit water allowed for analyzing the flexibility of each disaccharide ligand in the protein binding site. The higher affinity observed for binding to Gb3 epitope, both in vivo and on glycan array, is likely related to the presentation effect of the oligosaccharide on a surface, since only the Gb3 glycosphingolipid geometry is fully compatible with parallel insertion of neighboring trisaccharide heads in two binding sites of the same tetramer of PA-IL.
BMC Structural Biology, 2007
Background: Lectins are proteins of non-immune origin capable of binding saccharide structures with high specificity and affinity. Considering the high encoding capacity of oligosaccharides, this makes lectins important for adhesion and recognition. The present study is devoted to the PA-IIL lectin from Pseudomonas aeruginosa, an opportunistic human pathogen capable of causing lethal complications in cystic fibrosis patients. The lectin may play an important role in the process of virulence, recognizing specific saccharide structures and subsequently allowing the bacteria to adhere to the host cells. It displays high values of affinity towards monosaccharides, especially fucose -a feature caused by unusual binding mode, where two calcium ions participate in the interaction with saccharide. Investigating and understanding the nature of lectin-saccharide interactions holds a great potential of use in the field of drug design, namely the targeting and delivery of active compounds to the proper site of action.
ChemistrySelect, 2017
Due to the ability of Pseudomonas aeruginosa (PA) to develop antibiotic resistances, alternative therapeutic strategies have been proposed. Among others, carbohydrate multivalent molecules targeting lectin-based virulent factors have been widely reported in particular those targeting LecA. LecA is a tetravalent galactose specific lectin involved in biofilm formation and cell internalization. Herein, we report the synthesis of 36 galactoclusters built from galactosides with aromatic and non-aromatic aglycons and with an additional chain. The chains were either neutral or positively charged. Only the galactoclusters with naphthyl or tyrosine aglycon showed a moderate increase of binding for the positively charged 3-dimethylammonium propyl chain. In contrast, the non-aromatic galactoclusters display typically poorer binding properties towards LecA. The introduction of these side chains led to improved affinities up to becoming comparable to the high-affinity aromatic galactoclusters.
International Journal of Molecular Sciences
The Gram-negative bacterium Pseudomonas aeruginosa is an important opportunistic human pathogen associated with cystic fibrosis. P. aeruginosa produces two soluble lectins, the d-galactose-specific lectin PA-IL (LecA) and the l-fucose-specific lectin PA-IIL (LecB), among other virulence factors. These lectins play an important role in the adhesion to host cells and biofilm formation. Moreover, PA-IL is cytotoxic to respiratory cells in the primary culture. Therefore, these lectins are promising therapeutic targets. Specifically, carbohydrate-based compounds could inhibit their activity. In the present work, a 3-O-fucosyl lactose-containing tetravalent glycocluster was synthesized and utilized as a mutual ligand of galactophilic and fucophilic lectins. Pentaerythritol equipped with azido ethylene glycol-linkers was chosen as a multivalent scaffold and the glycocluster was constructed by coupling the scaffold with propargyl 3-O-fucosyl lactoside using an azide-alkyne 1,3-dipolar cyclo...