Gut Symbiont Secretes a Eukaryotic-Like Ubiquitin Protein That Mediates Intraspecies Antagonism.

Human gut species produce different types of toxins that antagonize closely related members of the gut microbiota. Some are toxic effectors delivered by type VI secretion systems, and others are non-contact-dependent secreted antimicrobial proteins. Many strains of secrete antimicrobial molecules, but only one of these toxins has been described to date ( secreted antimicrobial protein 1 [BSAP-1]).

Bacteroidales Secreted Antimicrobial Proteins Target Surface Molecules Necessary for Gut Colonization and Mediate Competition In Vivo.

Unlabelled: We recently showed that human gut Bacteroidales species secrete antimicrobial proteins (BSAPs), and we characterized in vitro the first such BSAP produced by Bacteroides fragilis In this study, we identified a second potent BSAP produced by the ubiquitous and abundant human gut species Bacteroides uniformis The two BSAPs contain a membrane attack complex/perforin (MACPF) domain but share very little sequence similarity. We identified the target molecules of BSAP-sensitive cells and showed that each BSAP targets a different class of surface molecule: BSAP-1 targets an outer membrane protein of sensitive B. fragilis strains, and BSAP-2 targets the O-antigen glycan of lipopolysaccharide (LPS) of sensitive B.

Type VI secretion systems of human gut Bacteroidales segregate into three genetic architectures, two of which are contained on mobile genetic elements.

Background: Type VI secretion systems (T6SSs) are contact-dependent antagonistic systems employed by Gram negative bacteria to intoxicate other bacteria or eukaryotic cells. T6SSs were recently discovered in a few Bacteroidetes strains, thereby extending the presence of these systems beyond Proteobacteria. The present study was designed to analyze in a global nature the diversity, abundance, and properties of T6SSs in the Bacteroidales, the most predominant Gram negative bacterial order of the human gut.

Systematic Identification of Cyclic-di-GMP Binding Proteins in Vibrio cholerae Reveals a Novel Class of Cyclic-di-GMP-Binding ATPases Associated with Type II Secretion Systems.

Cyclic-di-GMP (c-di-GMP) is a ubiquitous bacterial signaling molecule that regulates a variety of complex processes through a diverse set of c-di-GMP receptor proteins. We have utilized a systematic approach to identify c-di-GMP receptors from the pathogen Vibrio cholerae using the Differential Radial Capillary Action of Ligand Assay (DRaCALA). The DRaCALA screen identified a majority of known c-di-GMP binding proteins in V.

Systematic identification of conserved bacterial c-di-AMP receptor proteins.

Nucleotide signaling molecules are important messengers in key pathways that allow cellular responses to changing environments. Canonical secondary signaling molecules act through specific receptor proteins by direct binding to alter their activity. Cyclic diadenosine monophosphate (c-di-AMP) is an essential signaling molecule in bacteria that has only recently been discovered.

A rapid assay for affinity and kinetics of molecular interactions with nucleic acids.

The Differential Radial Capillary Action of Ligand Assay (DRaCALA) allows detection of protein interactions with low-molecular weight ligands based on separation of the protein-ligand complex by differential capillary action. Here, we present an application of DRaCALA to the study of nucleic acid-protein interactions using the Escherichia coli cyclic AMP receptor protein (CRP). CRP bound in DRaCALA specifically to (32)P-labeled oligonucleotides containing the consensus CRP binding site, but not to oligonucleotides with point mutations known to abrogate binding.

Differential radial capillary action of ligand assay for high-throughput detection of protein-metabolite interactions.

Interactions of proteins with low-molecular-weight ligands, such as metabolites, cofactors, and allosteric regulators, are important determinants of metabolism, gene regulation, and cellular homeostasis. Pharmaceuticals often target these interactions to interfere with regulatory pathways. We have developed a rapid, precise, and high-throughput method for quantitatively measuring protein-ligand interactions without the need to purify the protein when performed in cells with low background activity.