Many heterotrophic bacteria have the ability to make polyhedral structures containing metabolic enzymes that are bounded by a unilamellar protein shell (metabolosomes or enterosomes). These bacterial organelles contain enzymes associated with a specific metabolic process (e.g. 1,2-propanediol or ethanolamine utilization). We show that the 21 gene regulon specifying the pdu organelle and propanediol utilization enzymes from Citrobacter freundii is fully functional when cloned in Escherichia coli, both producing metabolosomes and allowing propanediol utilization. Genetic manipulation of the level of specific shell proteins resulted in the formation of aberrantly shaped metabolosomes, providing evidence for their involvement as delimiting entities in the organelle. This is the first demonstration of complete recombinant metabolosome activity transferred in a single step and supports phylogenetic evidence that the pdu genes are readily horizontally transmissible. One of the predicted shell proteins (PduT) was found to have a novel Fe-S center formed between four protein subunits. The recombinant model will facilitate future experiments establishing the structure and assembly of these multiprotein assemblages and their fate when the specific metabolic function is no longer required.It has been recognized for more than 30 years that all cyanobacteria (1) and some other chemoautotrophic bacteria (2) contain carboxysomes. These polyhedral cellular inclusions consist of a proteinaceous shell enclosing an active enzyme, ribulose bisphosphate carboxylase/oxygenase (RuBisCO). 4Their function is to enhance the fixation of carbon dioxide (3), a reaction of planetary significance in that marine cyanobacteria are responsible for the majority of global carbon fixation (4, 5). More recently, sequence similarity was noticed between carboxysome shell genes and metabolic operon genes associated with propanediol utilization (pdu) and ethanolamine utilization (eut) in a variety of heterotrophic bacteria found in the mammalian gut (3) and the environment. In growth conditions that induce these metabolic operons, polyhedral organelles resembling carboxysomes were observed on electron microscopy of Salmonella enterica serovar Typhimurium (6), Klebsiella oxytoca, Citrobacter freundii, and Escherichia coli (7). Bioinformatics analysis also locates genes resembling carboxysome shell genes in metabolic operons in Clostridium perfringens (8), Clostridium tetani (9), Listeria monocytogenes and Listeria innocua (10), Enterococcus faecalis (11), Lactobacillus collinoides (12), Citrobacter rodentium, 5 and Yersinia enterocolitica (13) among other organisms. The non-carboxysome polyhedral structures have been referred to as enterosomes (3) or metabolosomes (14), emphasizing their role in cellular metabolism.There is some considerable interest in how these proteinaceous organelles form and the arrangement of protein subunits that give rise to these remarkable macromolecular assemblies. In carboxysomes, there are thought to be a number of shell prot...
SummaryPseudomonas aeruginosa chronically colonizing the lungs of cystic fibrosis (CF) patients undergoes fast evolution leading to clonal divergence. More than half of the genotypes of P. aeruginosa clone C isolates exclusively from CF lung infection exhibit large chromosomal inversions (LCIs). To analyse the impact of LCIs, as a novel mechanism of bacterial adaptation, the underlying molecular mechanism was examined. Analysis of inversion breakpoints suggested an IS 6100 -induced coupled insertion-inversion mechanism. A selective advantage was created by insertion of IS 6100 into wbpM , pilB and mutS which leads to common CF phenotypes such as O-antigen and type IV pili deficiency and hypermutability. Speciation in bacteria is accompanied by LCIs. Therefore adaptation by LCIs that allows persistence of P. aeruginosa in the CF lung and species diversification in that new ecological niche can serve as a model for bacterial genome evolution.
A collection of 77 epidemiologically unrelated Pseudomonas aeruginosa isolates was screened for the occurrence of clone C isolates by the appearance of characteristic SpeI fragment patterns obtained by pulsed-field gel electrophoresis. Three strains with a clone C characteristic SpeI fragment pattern were found which also harbored the clone C-specific plasmid either in the free form or chromosomally integrated. Genomic islands were detected in the new clone C strains, as in already characterized clone C strains. Clone C not only infected cystic fibrosis patients throughout Europe, but was also found in the UK as an isolate in urinary tract infections and in peritoneal dialysis fluid, in addition to an otitis media isolate. Therefore, P. aeruginosa clone C is widely distributed in Europe, with a broad pathogenic potential.
The artificial sputum medium (ASM) was formulated to mimic the sputum of cystic fibrosis (CF) patients. The intra-and inter-patient variation in the composition of highly viscous CF sputum complicates the achievement of a reproducible culture condition to study microbial biofilms, for example, Pseudomonas aeruginosa colonization in the CF lung. However, the ASM is a homogenous non-viscous medium, which can be prepared in bulk quantities using a fixed set of ingredients. The conventional biofilm models always had cells attached to a solid biotic or abiotic surface submerged in a steady-state or continuous flow of culture medium. In contrast, P. aeruginosa grows in the CF lung under micro-aerophilic to anaerobic conditions in the form of microcolonies eventually forming a macro-aggregate/colony in which bacteria adhere to each other and to sputum components. Similarly, in ASM, P. aeruginosa forms micro-and macro-colonies and showed various phenotypes specific to CF isolates.
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