Abstract
Bacterial surfaces are decorated with carbohydrate structures that may serve as ligands for host receptors. Based on their ability to recognize specific sugar epitopes, plant lectins are extensively used for bacteria typing. We previously observed that the galactose-specific agglutinins from Ricinus communis (RCA) and Viscum album (VAA) exhibited differential binding to nontypeable Haemophilus influenzae (NTHi) clinical isolates, their binding being distinctly affected by truncation of the lipooligosaccharide (LOS). Here, we examined their binding to the structurally similar LOS molecules isolated from strains NTHi375 and RdKW20, using microarray binding assays, saturation transfer difference NMR, and molecular dynamics simulations. RCA bound the LOSRdKW20 glycoform displaying terminal Galβ(1,4)Glcβ, whereas VAA recognized the Galα(1,4)Galβ(1,4)Glcβ epitope in LOSNTHi375 but not in LOSRdKW20, unveiling a different presentation. Binding assays to whole bacterial cells were consistent with LOSNTHi375 serving as ligand for VAA, and also suggested recognition of the glycoprotein HMW1. Regarding RCA, comparable binding to NTHi375 and RdKW20 cells was observed. Interestingly, an increase in LOSNTHi375 abundance or expression of HMW1 in RdKW20 impaired RCA binding. Overall, the results revealed that, besides the LOS, other carbohydrate structures on the bacterial surface serve as lectin ligands, and highlighted the impact of the specific display of cell surface components on lectin binding.
Highlights
Ubiquitous in nature, carbohydrates mediate a myriad of recognition events, both in health and disease
The absence of this epitope in the NTHi375Δlic2A mutant, lacking the glycosyltransferase that adds β-galactose (Gal I) to the glucose residue linked to Hep III (Glc II), resulted in decreased binding of VAA compared to the wild type (WT) strain, indicating that the LOS may serve as docking point for this lectin
In this work we comparatively examined the binding of VAA and RCA to the LOS from NTHi375 and from the capsule-deficient H. influenzae laboratory strain RdKW20 (Fig. 1b) whose major glycoform presents terminal Galβ(1,4)Glc[28] (Gal I-Glc II in Fig. 1a,b,c) that could potentially be recognized by the two lectins[29,30,31]
Summary
Ubiquitous in nature, carbohydrates mediate a myriad of recognition events, both in health and disease. We developed novel bacteria-based microarrays and quartz crystal microbalance (QCM) chips for the screening of bacterial glycosignatures, by testing the binding of a panel of lectins with diverse carbohydrate-binding specificities, and quantitative analysis of lectin−bacteria interactions[23,24,25] Using this combined approach, different lectin-binding fingerprints were observed for six clinical isolates of nontypeable (non-capsulated) H. influenzae (NTHi), consistent with the above mentioned inter-strain heterogeneity of the bacterium[24]. The LOS of this NTHi strain (Fig. 1a) is known to contain the Galα(1,4)Galβ epitope in the chain extension linked to the distal manno-heptose (Hep III) of the Hep trisaccharide inner core (Gal II-Gal I in Fig. 1a,b,c)[26,27] The absence of this epitope in the NTHi375Δlic2A mutant, lacking the glycosyltransferase that adds β-galactose (Gal I) to the glucose residue linked to Hep III (Glc II), resulted in decreased binding of VAA compared to the wild type (WT) strain, indicating that the LOS may serve as docking point for this lectin. The results evidenced that, besides the LOS molecule, other carbohydrate structures on the bacterial surface are recognized by these two lectins, highlighting the complexity of lectin–bacteria interplays
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