RESUMEN
Aim: This study is mainly devoted to determining the ability of ∆FN3.1 protein fragments of Bifidobacterium (B.) longum subsp. longum GT15, namely two FN3 domains (2D FN3) and a C-terminal domain (CD FN3), to bind to tumor necrosis factor-alpha (TNF-α). Methods: Fragments of the fn3 gene encoding the 2D FN3 and CD FN3 were cloned in Escherichia (E.) coli. In order to assess the binding specificity between 2D FN3 and CD FN3 to TNFα, we employed the previously developed sandwich ELISA system to detect any specific interactions between the purified protein and any of the studied cytokines. The trRosetta software was used to build 3D models of the ∆FN3.1, 2D FN3, and CD FN3 proteins. The detection of polymorphism in the amino acid sequences of the studied proteins and the analysis of human gut-derived bacterial proteins carrying FN3 domains were performed in silico. Results: We experimentally showed that neither 2D FN3 nor CD FN3 alone can bind to TNFα. Prediction of the 3D structures of ΔFN3.1, 2D FN3, and CD FN3 suggested that only ΔFN3.1 can form a pocket allowing binding with TNFα to occur. Polymorphism analysis of amino acid sequences of ΔFN3.1 proteins in B. longum strains uncovered substitutions that can alter the conformation of the spatial structure of the ΔFN3.1 protein. We also analyzed human gut-derived bacterial proteins harboring FN3 domains which allowed us to differentiate between those containing motifs of cytokine receptors (MCRs) in their FN3 domains and those lacking them. Conclusion: Only the complete ∆FN3.1 protein can selectively bind to TNFα. Analysis of 3D models of the 2D FN3, CD FN3, and ΔFN3.1 proteins showed that only the ΔFN3.1 protein is potentially capable of forming a pocket allowing TNFα binding to occur. Only FN3 domains containing MCRs exhibited sequence homology with FN3 domains of human proteins.
RESUMEN
Bifidobacteria are some of the major agents that shaped the immune system of many members of the animal kingdom during their evolution. Over recent years, the question of concrete mechanisms underlying the immunomodulatory properties of bifidobacteria has been addressed in both animal and human studies. A possible candidate for this role has been discovered recently. The PFNA cluster, consisting of five core genes, pkb2, fn3, aaa-atp, duf58, tgm, has been found in all gut-dwelling autochthonous bifidobacterial species of humans. The sensory region of the species-specific serine-threonine protein kinase (PKB2), the transmembrane region of the microbial transglutaminase (TGM), and the type-III fibronectin domain-containing protein (FN3) encoded by the I gene imply that the PFNA cluster might be implicated in the interaction between bacteria and the host immune system. Moreover, the FN3 protein encoded by one of the genes making up the PFNA cluster, contains domains and motifs of cytokine receptors capable of selectively binding TNF-α. The PFNA cluster could play an important role for sensing signals of the immune system. Among the practical implications of this finding is the creation of anti-inflammatory drugs aimed at alleviating cytokine storms, one of the dire consequences resulting from SARS-CoV-2 infection.
Asunto(s)
Proteínas Bacterianas/genética , Bifidobacterium/fisiología , COVID-19/terapia , Proteínas Serina-Treonina Quinasas/genética , Proteínas Bacterianas/química , Proteínas Bacterianas/metabolismo , COVID-19/inmunología , COVID-19/virología , Síndrome de Liberación de Citoquinas/inmunología , Síndrome de Liberación de Citoquinas/prevención & control , Citocinas/química , Citocinas/metabolismo , Humanos , Sistema Inmunológico , Operón/genética , Proteínas Serina-Treonina Quinasas/química , Proteínas Serina-Treonina Quinasas/metabolismo , SARS-CoV-2/aislamiento & purificaciónRESUMEN
The draft genome sequences of Bifidobacterium angulatum GT102 and Bifidobacterium adolescentis 150 strains isolated from the human intestinal microbiota are reported. Both strains are able to produce gamma-aminobutyric acid (GABA). Detailed genomes analysis will help to understand the role of GABA in the functioning of gut-brain axis.
RESUMEN
Six genes encoding the bifidobacterial Hanks-type (eukaryote-like) serine/threonine protein kinases (STPK) were identified and classified. The genome of each bifidobacterial strain contains four conserved genes and one species-specific gene. Bifidobacterium longum and Bifidobacterium bifidum possess the unique gene found only in these species. The STPK genes of Russian industrial probiotic strain B. longum B379M were cloned and sequenced. The expression of these genes in Escherichia coli and bifidobacteria was observed. Autophosphorylation of the conserved STPK Pkb5 and species-specific STPK Pkb2 was demonstrated. This is the first report on Hanks-type STPK in bifidobacteria.