Whole Cell Cross-Linking to Discover Host–Microbe Protein Cognate Receptor/Ligand Pairs

Abstract

Bacterial surface ligands mediate interactions with the host cell during association that determines the specific outcome for the host–microbe association. The association begins with receptors on the host cell binding ligands on the microbial cell to form a partnership that initiates responses in both cells. Methods to determine the specific cognate partnerships are lacking. Determining these molecular interactions between the host and microbial surfaces are difficult, yet crucial in defining biologically important events that are triggered during association of the microbiome, and critical in defining the initiating signal from the host membrane that results in pathology or commensal association. In this study, we designed an approach to discover cognate host–microbe receptor/ligand pairs using a covalent cross-linking strategy with whole cells. Protein/protein cross-linking occurred when the interacting molecules were within 9–12 Å, allowing for identification of specific pairs of proteins from the host and microbe that define the molecular interaction during association. To validate the method three different bacteria with three previously known protein/protein partnerships were examined. The exact interactions were confirmed and led to discovery of additional partnerships that were not recognized as cognate partners, but were previously reported to be involved in bacterial interactions. Additionally, three unknown receptor/ligand partners were discovered and validated with in vitro infection assays by blocking the putative host receptor and deleting the bacterial ligand. Subsequently, Salmonella enterica sv. Typhimurium was cross-linked to differentiated colonic epithelial cells (caco-2) to discover four previously unknown host receptors bound to three previously undefined host ligands for Salmonella. This approach resulted in a priori discovery of previously unknown and biologically important molecules for host/microbe association that were casually reported to mediate bacterial invasion. The whole cell cross-linking approach promises to enable discovery of possible targets to modulate interaction of the microbiome with the host that are important in infection and commensalism, both of with initiate a host response.

Marine Microalgae Biomolecules and Their Adhesion Capacity to Salmonella enterica sv. Typhimurium

“…The Salmonella adhesion is mediated via protein-carbohydrate or a protein-protein interactions [57], thus proteins and glycosides (SQG and DGG) in alkaline fractions could be synergistically cooperating with polysaccharides binding to bacteria through other mechanisms. For Salmonella Typhimurium, the object of this study, protein-protein interactions are mediated by fimbrial and non-fimbrial adhesins and involve the extracellular matrix components of the host cell [58][59][60][61][62][63]. Several studies reported that milk and colostrum fractions containing proteins (β-lactoglobulin, α-lactalbumin) and glycopeptides (glycomacropeptide) show anti-adherence properties against Salmonella Typhimurium, Salmonella enteritidis, enteropathogenic Escherichia coli, and enterohemorrhagic E. coli, among others [11,[64][65][66].…”

MicrobioLink: An Integrated Computational Pipeline to Infer Functional Effects of Microbiome–Host Interactions

“…Microbiota-host interactions happen in almost every known organism, shaping their metabolism and evolution [1,2]. In many ecosystems, the microbiome plays an important role as manifested by its dynamic interactions with different hosts [3].…”

“…Besides the metabolite-mediated interactions, protein-protein interactions (PPIs) are one of the most relevant types of molecular interplay between microbes and host organisms [12]. Experimental techniques to test interspecies PPIs are time consuming and have limitations imposed by cost [1,[17][18][19][20][21]. Hence, there is a dearth of validated interspecies PPIs in publicly available databases, and these are mostly limited to pathogens.…”

MicrobioLink: An integrated computational pipeline to infer functional effects of microbiome-host interactions

“…Microbiota-host interactions happen in most organisms, shaping their metabolism and evolution [1][2][3]. In many ecosystems, microbial communities play important roles in the dynamic interactions with the host [4].…”

“…Next to metabolite mediated interactions, protein-protein interactions (PPIs) are one of the most relevant and reported types of molecular interactions between the host and microbes [17][18][19][20]. However, experimental techniques to probe inter-species PPIs are time consuming and have limitations imposed by cost [2,14,16,[22][23][24]. Hence, there is a dearth of validated inter-species PPIs in publicly available databases and are mostly limited to pathogens.…”