Use of Atomic Force Microscopy to Study the Multi-Modular Interaction of Bacterial Adhesins to Mucins

Abstract

The mucus layer covering the gastrointestinal (GI) epithelium is critical in selecting and maintaining homeostatic interactions with our gut bacteria. However, the molecular details of these interactions are not well understood. Here, we provide mechanistic insights into the adhesion properties of the canonical mucus-binding protein (MUB), a large multi-repeat cell–surface adhesin found in Lactobacillus inhabiting the GI tract. We used atomic force microscopy to unravel the mechanism driving MUB-mediated adhesion to mucins. Using single-molecule force spectroscopy we showed that MUB displayed remarkable adhesive properties favouring a nanospring-like adhesion model between MUB and mucin mediated by unfolding of the multiple repeats constituting the adhesin. We obtained direct evidence for MUB self-interaction; MUB–MUB followed a similar binding pattern, confirming that MUB modular structure mediated such mechanism. This was in marked contrast with the mucin adhesion behaviour presented by Galectin-3 (Gal-3), a mammalian lectin characterised by a single carbohydrate binding domain (CRD). The binding mechanisms reported here perfectly match the particular structural organization of MUB, which maximizes interactions with the mucin glycan receptors through its long and linear multi-repeat structure, potentiating the retention of bacteria within the outer mucus layer.

Unravelling the specificity and mechanism of sialic acid recognition by the gut symbiont Ruminococcus gnavus

“…Although a wide number of microbial lectins have been functionally and structurally characterised to date, especially from pathogens, only a few carbohydrate-binding proteins present in gut bacteria which interact with mucus have been structurally characterised 13 , 15 . Interactions between bacterial adhesins from gut commensals and mucin glycans are generally of low affinity, in line with the localisation of these bacteria within the outer mucus layer 48 , 49 . Here we showed that Rg CBM40 could recognise mucins with binding affinity increasing with sialic acid level.…”

Surface Proteins of Lactococcus lactis: Bacterial Resources for Muco-adhesion in the Gastrointestinal Tract

“…Contrary to the MucBP domain also present in pathogens ( Popowska et al, 2017 ), the Mub domain is predominantly found in LAB and is highly abundant in lactobacilli of the gut microbiota ( Boekhorst et al, 2006 ). The best characterized MUB protein from Lactobacillus reuteri endowed with 14 Mub domains ( Roos and Jonsson, 2002 ) binds mucus via multiple interactions involving terminal sialylated mucin glycans ( Etzold and Juge, 2014 ; Gunning et al, 2016 ). In L. lactis plasmid free laboratory strains IL1403 and MG1363, only one MUB protein with four Mub domains and devoid of signal sequence, is encoded in the chromosome sequence ( Boekhorst et al, 2006 ).…”

Probiotic Lactobacillus reuteri DSM 17938: what is known about it today?

“…Еще один из возможных механизмов адгезии -это связывание поверхностных молекул бактерий со слоем слизи. Среди штаммов L. reuteri наибольшее разнообразие слизесвязывающих (MUB) и MUB-подобных белков кодируется Lactobacillalus-специфическими кластерами генов (адгезинами) [11][12][13]. В исследованиях показано, что разнообразие MUBs среди штаммов L. reuteri и различия в численности MUB на клеточной поверхности значительно коррелируют с их способностью связывать слизь [14].…”