Abstract
Brucella species are Gram-negative, facultative intracellular pathogens responsible for a worldwide zoonosis. The envelope of Brucella exhibits unique characteristics that make these bacteria furtive pathogens and resistant to several host defence compounds. We have identified a Brucella suis gene (mapB) that appeared to be crucial for cell envelope integrity. Indeed, the typical resistance of Brucella to both lysozyme and the cationic lipopeptide polymyxin B was markedly reduced in a ∆mapB mutant. MapB turned out to represent a TamB orthologue. This last protein, together with TamA, a protein belonging to the Omp85 family, form a complex that has been proposed to participate in the translocation of autotransporter proteins across the outer membrane (OM). Accordingly, we observed that MapB is required for proper assembly of an autotransporter adhesin in the OM, as most of the autotransporter accumulated in the mutant cell periplasm. Both assessment of the relative amounts of other specific outer membrane proteins (OMPs) and a proteome approach indicated that the absence of MapB did not lead to an extensive alteration in OMP abundance, but to a reduction in the relative amounts of a protein subset, including proteins from the Omp25/31 family. Electron microscopy revealed that ∆mapB cells exhibit multiple anomalies in cell morphology, indicating that the absence of the TamB homologue in B. suis severely affects cell division. Finally, ∆mapB cells were impaired in macrophage infection and showed an attenuated virulence phenotype in the mouse model. Collectively, our results indicate that the role of B. suis TamB homologue is not restricted to participating in the translocation of autotransporters across the OM but that it is essential for OM stability and protein composition and that it is involved in cell envelope biogenesis, a process that is inherently coordinated with cell division.
Highlights
Bacteria of the genus Brucella are gram-negative bacteria, responsible for brucellosis, a disease characterized by chronic infections, abortions and infertility in animals, and chronic fatigue in humans[1]
The Translocation and Assembly Module (TAM) system consists of TamA, another member of the Omp[85] superfamily, and TamB, a large protein that is inserted in the inner membrane (IM) by an N-terminal non-cleavable signal peptide; the rest of TamB exhibits an extensive β-helical structure that is immersed in the periplasm[19,20]
While performed an in silico analysis to identify Brucella autotransporters the BR0049 gene from B. suis 1330 came out as a possible adhesin with a low similarity to autotransporters. This was due to the abundance of β-helix strands that are predicted along almost the entire protein and to a β-sheet structure found in the very C-terminus of BR0049 and its orthologues from other Alphaproteobacteria[21]
Summary
Bacteria of the genus Brucella are gram-negative bacteria, responsible for brucellosis, a disease characterized by chronic infections, abortions and infertility in animals, and chronic fatigue in humans[1]. Proteins from the TamB family were found to be widely distributed in most of the gram-negative bacterial lineages, and surprisingly, even in numerous genera that do not harbor autotransporters[21] This raises the question of which is the primary role of TamB in the biogenesis of the bacterial cell envelope. While performed an in silico analysis to identify Brucella autotransporters the BR0049 gene from B. suis 1330 came out as a possible adhesin with a low similarity to autotransporters This was due to the abundance of β-helix strands that are predicted along almost the entire protein and to a β-sheet structure found in the very C-terminus of BR0049 and its orthologues from other Alphaproteobacteria[21]. We propose that BR0049 is involved in cell envelope biogenesis, in a process that is inherently coordinated with cellular division and that is crucial for cellular integrity
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