RESUMEN
Bacteria in the gastrointestinal tract produce amino acid bile acid amidates that can affect host-mediated metabolic processes1-6; however, the bacterial gene(s) responsible for their production remain unknown. Herein, we report that bile salt hydrolase (BSH) possesses dual functions in bile acid metabolism. Specifically, we identified a previously unknown role for BSH as an amine N-acyltransferase that conjugates amines to bile acids, thus forming bacterial bile acid amidates (BBAAs). To characterize this amine N-acyltransferase BSH activity, we used pharmacological inhibition of BSH, heterologous expression of bsh and mutants in Escherichia coli and bsh knockout and complementation in Bacteroides fragilis to demonstrate that BSH generates BBAAs. We further show in a human infant cohort that BBAA production is positively correlated with the colonization of bsh-expressing bacteria. Lastly, we report that in cell culture models, BBAAs activate host ligand-activated transcription factors including the pregnane X receptor and the aryl hydrocarbon receptor. These findings enhance our understanding of how gut bacteria, through the promiscuous actions of BSH, have a significant role in regulating the bile acid metabolic network.
Asunto(s)
Aciltransferasas , Amidohidrolasas , Aminas , Ácidos y Sales Biliares , Biocatálisis , Microbioma Gastrointestinal , Humanos , Aciltransferasas/metabolismo , Amidohidrolasas/metabolismo , Aminas/química , Aminas/metabolismo , Bacteroides fragilis/enzimología , Bacteroides fragilis/genética , Bacteroides fragilis/metabolismo , Ácidos y Sales Biliares/química , Ácidos y Sales Biliares/metabolismo , Estudios de Cohortes , Escherichia coli/enzimología , Escherichia coli/genética , Escherichia coli/metabolismo , Microbioma Gastrointestinal/fisiología , Ligandos , Receptor X de Pregnano/metabolismo , Receptores de Hidrocarburo de Aril/metabolismo , Factores de Transcripción/metabolismo , Lactante , Técnicas de Cultivo de CélulaRESUMEN
Heme-based sensor proteins are used by organisms to control signaling and physiological effects in response to their gaseous environment. Globin-coupled sensors (GCS) are oxygen-sensing proteins that are widely distributed in bacteria. These proteins consist of a heme globin domain linked by a middle domain to various output domains, including diguanylate cyclase domains, which are responsible for synthesizing c-di-GMP, a bacterial second messenger crucial for regulating biofilm formation. To understand the roles of heme pocket residues in controlling activity of the diguanylate cyclase domain, variants of the Pectobacterium carotovorum GCS (PccGCS) were characterized by enzyme kinetics and resonance Raman (rR) spectroscopy. Results of these studies have identified roles for hydrogen bonding and heme edge residues in modulating heme pocket conformation and flexibility. Better understanding of the ligand-dependent GCS signaling mechanism and the residues involved may allow for future development of methods to control O2-dependent c-di-GMP production.
Asunto(s)
Proteínas Bacterianas , Hemo , Enlace de Hidrógeno , Pectobacterium carotovorum , Liasas de Fósforo-Oxígeno , Espectrometría Raman , Liasas de Fósforo-Oxígeno/metabolismo , Liasas de Fósforo-Oxígeno/química , Espectrometría Raman/métodos , Hemo/química , Hemo/metabolismo , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , Pectobacterium carotovorum/enzimología , Globinas/química , Globinas/metabolismo , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , GMP Cíclico/química , Proteínas de Escherichia coliRESUMEN
Bacteria sense and respond to their environment, allowing them to maximize their survival and growth under changing conditions, such as oxygen levels. Direct oxygen-sensing proteins allow bacteria to rapidly sense concentration changes and adapt by regulating signaling pathways and/or cellular machinery. Recent work has identified roles for direct oxygen-sensing proteins in controlling second messenger levels and motility machinery, as well as effects on biofilm formation, virulence, and motility. In this review, we discuss recent progress in understanding O2-dependent regulation of cyclic di-GMP signaling and motility and highlight the emerging importance in controlling bacterial physiology and behavior.
Asunto(s)
Proteínas de Escherichia coli , Hemoproteínas , GMP Cíclico/metabolismo , Hemoproteínas/genética , Hemoproteínas/metabolismo , Oxígeno/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Biopelículas , Sistemas de Mensajero Secundario/fisiología , Bacterias/genética , Bacterias/metabolismo , Proteínas de Escherichia coli/genética , Hemo/metabolismo , Regulación Bacteriana de la Expresión GénicaRESUMEN
Bacteria sense and respond to gaseous ligand changes in the environment to regulate a multitude of behaviors, including the production of the secondary messengers cyclic di-GMP. Gas sensing can be difficult to measure due to the high concentration of the oxygen in the atmosphere, particularly in redox-sensitive systems. Here, we describe a method for anaerobic quantification of cyclic di-GMP production which can be used to measure the impact of molecular oxygen, nitric oxide, and carbon monoxide on the catalysis of a diguanylate cyclase-containing protein and the possible pitfalls in the experimental procedure.