Exploration of Galectin Ligands Displayed on Gram-Negative Respiratory Bacterial Pathogens with Different Cell Surface Architectures

Abstract

Galectins bind various pathogens through recognition of distinct carbohydrate structures. In this work, we examined the binding of four human galectins to the Gram-negative bacteria Klebsiella pneumoniae (Kpn) and non-typeable Haemophilus influenzae (NTHi), which display different surface glycans. In particular, Kpn cells are covered by a polysaccharide capsule and display an O-chain-containing lipopolysaccharide (LPS), whereas NTHi is not capsulated and its LPS, termed lipooligosacccharide (LOS), does not contain O-chain. Binding assays to microarray-printed bacteria revealed that galectins-3, -4, and -8, but not galectin-1, bind to Kpn and NTHi cells, and confocal microscopy attested binding to bacterial cells in suspension. The three galectins bound to array-printed Kpn LPS. Moreover, analysis of galectin binding to mutant Kpn cells evidenced that the O-chain is the docking point for galectins on wild type Kpn. Galectins-3, -4, and -8 also bound the NTHi LOS. Microarray-assisted comparison of the binding to full-length and truncated LOSs, as well as to wild type and mutant cells, supported LOS involvement in galectin binding to NTHi. However, deletion of the entire LOS oligosaccharide chain actually increased binding to NTHi cells, indicating the availability of other ligands on the bacterial surface, as similarly inferred for Kpn cells devoid of both O-chain and capsule. Altogether, the results illustrate galectins’ versatility for recognizing different bacterial structures, and point out the occurrence of so far overlooked galectin ligands on bacterial surfaces.

Whole microbe arrays accurately predict interactions and overall antimicrobial activity of galectin-8 toward distinct strains of Streptococcus pneumoniae

“…The Galβ1-3Gal linkage may ultimately be responsible for supporting glycan binding, but it is not clear to what extent the Galβ1-2 linkage may likewise influence Gal-8 binding to this microbe 75 . Importantly, Gal-8 also bound Haemophilus influenza, consistent with recent findings 26 , 31 . As noted, the synthetic complexity of microbial glycans prohibits a detailed analysis of the unique glycan determinants that are ultimately responsible for dictating differences in binding affinity observed by Gal-8 toward unique microbial glycans.…”

“…Similar to Gal-8N analysis on the MGM, Gal-8N likewise failed to display appreciable binding on the S. pneumonia MMA (Fig. 5 b), but did recognize Haemophilus influenzae (HI), which served as positive control for Gal-8 and Gal-8N 31 . In contrast to results obtained following flow cytometric examination of Gal-8C toward Sp 14, Gal-8C failed to recognize Sp 14 in the MMA format, although it did bind to Sp 33F, largely in agreement with MGM findings.…”

Lipopolysaccharide O-antigen molecular and supramolecular modifications of plant root microbiota are pivotal for host recognition

“…Herbaspirillum Root189 LPS was printed as triplicates at six different concentrations (from 1 to 0.003 mg ml -1 ) on 16-pad nitrocellulose-coated glass slides (Grace Biolabs ONCYTE NOVA) using a non-contact arrayer (Sprint, Arrayjet Ltd.) (Campanero-Rhodes, 2021). Control (glyco)proteins (fetuin, asialofetuin, ribonuclease B, and ribonuclease A) were printed in parallel at concentrations ranging from 1 to 0.03 mg ml -1 .…”

Galectins: Their Network and Roles in Infection/Immunity/Tumor Growth Control 2021

“…Campanero-Rhodes et al examined the binding of four human galectins to the Gram-negative bacteria [ 16 ]. Galectins have been shown to bind to the surfaces of some pathogens and products released by the pathogens [ 17 ].…”