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Biochimica Et Biophysica Acta Aug 2014Outer membrane vesicles (OMVs) are constitutively produced by all Gram-negative bacteria. OMVs form when buds from the outer membrane (OM) of cells encapsulate... (Review)
Review
Outer membrane vesicles (OMVs) are constitutively produced by all Gram-negative bacteria. OMVs form when buds from the outer membrane (OM) of cells encapsulate periplasmic material and pinch off from the OM to form spheroid particles approximately 10 to 300nm in diameter. OMVs accomplish a diversity of functional roles yet the OMV's utility is ultimately determined by its unique composition. Inclusion into OMVs may impart a variety of benefits to the protein cargo, including: protection from proteolytic degradation, enhancement of long-distance delivery, specificity in host-cell targeting, modulation of the immune response, coordinated secretion with other bacterial effectors, and/or exposure to a unique function-promoting environment. Many enriched OMV-associated components are virulence factors, aiding in host cell destruction, immune system evasion, host cell invasion, or antibiotic resistance. Although the mechanistic details of how proteins become enriched as OMV cargo remain elusive, recent data on OM biogenesis and relationships between LPS structure and OMV-cargo inclusion rates shed light on potential models for OM organization and consequent OMV budding. In this review, mechanisms based on pre-existing OM microdomains are proposed to explain how cargo may experience differing levels of enrichment in OMVs and degrees of association with OMVs during extracellular export. This article is part of a Special Issue entitled: Protein trafficking and secretion in bacteria. Guest Editors: Anastassios Economou and Ross Dalbey.
Topics: Bacterial Outer Membrane Proteins; Gram-Negative Bacteria; Periplasm; Periplasmic Proteins; Protein Transport; Transport Vesicles; Virulence Factors
PubMed: 24370777
DOI: 10.1016/j.bbamcr.2013.12.011 -
MSphere Dec 2020Two-component systems control periplasmic Cu homeostasis in Gram-negative bacteria. In characterized systems such as CusRS, upon Cu binding to the periplasmic sensing...
Two-component systems control periplasmic Cu homeostasis in Gram-negative bacteria. In characterized systems such as CusRS, upon Cu binding to the periplasmic sensing region of CusS, a cytoplasmic phosphotransfer domain of the sensor phosphorylates the response regulator CusR. This drives the expression of efflux transporters, chaperones, and redox enzymes to ameliorate metal toxic effects. Here, we show that the two-component sensor histidine kinase CopS exhibits a Cu-dependent phosphatase activity that maintains CopR in a nonphosphorylated state when the periplasmic Cu levels are below the activation threshold of CopS. Upon Cu binding to the sensor, the phosphatase activity is blocked and the phosphorylated CopR activates transcription of the CopRS regulon. Supporting the model, mutagenesis experiments revealed that the Δ strain exhibits maximal expression of the CopRS regulon, lower intracellular Cu levels, and increased Cu tolerance compared to wild-type cells. The invariant phosphoacceptor residue His of CopS was not required for the phosphatase activity itself but was necessary for its Cu dependency. To sense the metal, the periplasmic domain of CopS binds two Cu ions at its dimeric interface. Homology modeling of CopS based on CusS structure (four Ag binding sites) clearly supports the different binding stoichiometries in the two systems. Interestingly, CopS binds Cu with 3 × 10 M affinity, pointing to the absence of free (hydrated) Cu in the periplasm. Copper is a micronutrient required as cofactor in redox enzymes. When free, copper is toxic, mismetallating proteins and generating damaging free radicals. Consequently, copper overload is a strategy that eukaryotic cells use to combat pathogens. Bacteria have developed copper-sensing transcription factors to control copper homeostasis. The cell envelope is the first compartment that has to cope with copper stress. Dedicated two-component systems control the periplasmic response to metal overload. This paper shows that the sensor kinase of the copper-sensing two-component system present in exhibits a signal-dependent phosphatase activity controlling the activation of its cognate response regulator, distinct from previously described periplasmic Cu sensors. Importantly, the data show that the system is activated by copper levels compatible with the absence of free copper in the cell periplasm. These observations emphasize the diversity of molecular mechanisms that have evolved in bacteria to manage the copper cellular distribution.
Topics: Bacterial Proteins; Biological Transport; Cell Membrane; Copper; Escherichia coli; Histidine Kinase; Homeostasis; Periplasm; Phosphoric Monoester Hydrolases; Phosphorylation; Pseudomonas aeruginosa; Regulon
PubMed: 33361129
DOI: 10.1128/mSphere.01193-20 -
Current Opinion in Structural Biology Aug 2008The cell envelope of Gram-negative bacteria protects the organism from environmental stresses, components of the innate immune response, and the actions of other... (Review)
Review
The cell envelope of Gram-negative bacteria protects the organism from environmental stresses, components of the innate immune response, and the actions of other antagonistic molecules. However, the complexity of the cell envelope dictated by these protective roles creates a significant challenge for assembly of the outer membrane. Extensive research has focused on the export and assembly of outer membrane proteins and there is continuing progress in this area. By contrast, knowledge of the export and assembly of complex glycoconjugates in the outer membrane has been limited until recently. New structural and biochemical information identifies an envelope-spanning molecular scaffold for the export of group 1 capsular polysaccharides and provides insight into a complex molecular machine.
Topics: Bacterial Outer Membrane Proteins; Gram-Negative Bacteria; Lipopolysaccharides; Lipoproteins; Models, Molecular; Periplasm; Protein Conformation; Protein Folding
PubMed: 18495473
DOI: 10.1016/j.sbi.2008.04.001 -
Annual Review of Microbiology 2001Envelope stress responses play important physiological roles in a variety of processes, including protein folding, cell wall biosynthesis, and pathogenesis. Many of... (Review)
Review
Envelope stress responses play important physiological roles in a variety of processes, including protein folding, cell wall biosynthesis, and pathogenesis. Many of these responses are controlled by extracytoplasmic function (ECF) sigma factors that respond to external signals by means of a membrane-localized anti-sigma factor. One of the best-characterized, ECF-regulated responses is the sigma(E) envelope stress response of Escherichia coli. The sigma(E) pathway ensures proper assembly of outer-membrane proteins (OMP) by controlling expression of genes involved in OMP folding and degradation in response to envelope stresses that disrupt these processes. Prevailing evidence suggests that, in E. coli, a second envelope stress response controlled by the Cpx two-component system ensures proper pilus assembly. The sensor kinase CpxA recognizes misfolded periplasmic proteins, such as those generated during pilus assembly, and transduces this signal to the response regulator CpxR through conserved phosphotransfer reactions. Phosphorylated CpxR activates transcription of periplasmic factors necessary for pilus assembly.
Topics: Bacterial Outer Membrane Proteins; Bacterial Physiological Phenomena; Bacterial Proteins; Escherichia coli Proteins; Fimbriae, Bacterial; Heat-Shock Proteins; Periplasm; Protein Folding; Protein Kinases; Sigma Factor; Signal Transduction
PubMed: 11544368
DOI: 10.1146/annurev.micro.55.1.591 -
International Journal of Medical... Feb 2020The occurrence of antibiotic resistance bacteria has become a major threat to public health. We have recently discovered a transcriptional activator that belongs to MarR...
The occurrence of antibiotic resistance bacteria has become a major threat to public health. We have recently discovered a transcriptional activator that belongs to MarR family, EstR, and an esterase B (EstB) with a newly proposed de-arenethiolase activity from Sphingobium sp. SM42. De-arenethiolase activity involves the removal of the small aromatic side chain of cephalosporin antibiotics as an excellent leaving group by the enzymatic CS bond cleavage. Here, we report the regulation of estB through EstR as an activator in response to a third generation cephalosporin, cefoperazone, antibiotic. Cefoperazone induced the expression of estB in wild type Sphingobium sp., but not in the estR knockout strain, and the induction was restored in the complemented strain. Moreover, we revealed the importance of EstB localization in periplasm. Since EsB has the ability to inactivate selected β-lactam antibiotics in vitro, it is possible that the enzyme works at the periplasmic space of Gram negative bacteria similar to β-lactamases. EstB was genetically engineered by incorporating NlpA binding motif, or OmpA signal sequence, or SpyTag-SpyCatcher to the estB gene to mobilize it to different compartments of periplasm; inner membrane, outer membrane, and periplasmic space, respectively. Surprisingly, we found that Sphingobium sp. SM42 and E. coli expressing EstB at the periplasm were more sensitive to cefoperazone. The possible drug enhancement mechanism by enzyme was proposed. This work might lead to a novel strategy to tackle antibiotic resistance problem.
Topics: Cefoperazone; Cephalosporins; Escherichia coli; Gene Expression Regulation, Bacterial; Periplasm; Protein Sorting Signals; Serine Endopeptidases; Sphingomonadaceae; Transcription Factors
PubMed: 32005588
DOI: 10.1016/j.ijmm.2020.151396 -
Applied Biochemistry and Biotechnology Oct 2017Molecular chaperones and protein folding factors of bacterial periplasmic space play important roles in assisting disulfide bond formation and proper protein folding. In...
Molecular chaperones and protein folding factors of bacterial periplasmic space play important roles in assisting disulfide bond formation and proper protein folding. In this study, effects of disulfide bond protein (Dsb) families were investigated on assembly of 3F3 Fab, an antibody inhibitor targeting matrix metalloproteinase-14 (MMP-14). By optimizing DsbA/C co-expression, promoter for 3F3 Fab, host strains, and culture media and conditions, a high yield of 30-mg purified 3F3 Fab per liter culture was achieved. Produced 3F3 Fab exhibited binding affinity of 34 nM and inhibition potency of 970 nM. This established method of DsbA/C co-expression can be applied to produce other important disulfide bond-dependent recombinant proteins in E. coli periplasm.
Topics: Escherichia coli; Escherichia coli Proteins; Humans; Immunoglobulin Fab Fragments; Periplasm; Protein Disulfide-Isomerases; Protein Folding; Recombinant Proteins
PubMed: 28488120
DOI: 10.1007/s12010-017-2502-8 -
Sequential closure of the cytoplasm and then the periplasm during cell division in Escherichia coli.Journal of Bacteriology Feb 2012To visualize the latter stages of cell division in live Escherichia coli, we have carried out fluorescence recovery after photobleaching (FRAP) on 121 cells expressing...
To visualize the latter stages of cell division in live Escherichia coli, we have carried out fluorescence recovery after photobleaching (FRAP) on 121 cells expressing cytoplasmic green fluorescent protein and periplasmic mCherry. Our data show conclusively that the cytoplasm is sealed prior to the periplasm during the division event.
Topics: Cell Division; Cytoplasm; Escherichia coli; Escherichia coli Proteins; Fluorescence Recovery After Photobleaching; Green Fluorescent Proteins; Periplasm; Recombinant Fusion Proteins
PubMed: 22101847
DOI: 10.1128/JB.06091-11 -
Scientific Reports Nov 2016Carbohydrate polymers are industrially and medically important. For instance, a polysaccharide, alginate (from seaweed), is widely used in food, textile and...
Carbohydrate polymers are industrially and medically important. For instance, a polysaccharide, alginate (from seaweed), is widely used in food, textile and pharmaceutical industries. Certain bacteria also produce alginate through membrane spanning multi-protein complexes. Using Pseudomonas aeruginosa as a model organism, we investigated the biological function of an alginate degrading enzyme, AlgL, in alginate production and biofilm formation. We showed that AlgL negatively impacts alginate production through its enzymatic activity. We also demonstrated that deletion of AlgL does not interfere with polymer length control, epimerization degree or stability of the biosynthesis complex, arguing that AlgL is a free periplasmic protein dispensable for alginate production. This was further supported by our protein-stability and interaction experiments. Interestingly, over-production of AlgL interfered with polymer length control, suggesting that AlgL could be loosely associated with the biosynthesis complex. In addition, chromosomal expression of algL enhanced alginate O-acetylation; both attachment and dispersal stages of the bacterial biofilm lifecycle were sensitive to the level of O-acetylation. Since this modification also protects the pathogen against host defences and enhances other virulence factors, chromosomal expression of algL could be important for the pathogenicity of this organism. Overall, this work improves our understanding of bacterial alginate production and provides new knowledge for alginate production and disease control.
Topics: Acetylation; Alginates; Bacterial Proteins; Biofilms; Gene Deletion; Glucuronic Acid; Hexuronic Acids; Periplasm; Polysaccharide-Lyases; Protein Stability; Pseudomonas aeruginosa
PubMed: 27824067
DOI: 10.1038/srep31249 -
Pyoverdine biosynthesis and secretion in Pseudomonas aeruginosa: implications for metal homeostasis.Environmental Microbiology Jun 2013Pyoverdines are siderophores produced by fluorescent Pseudomonads to acquire iron. At least 60 different pyoverdines produced by diverse strains have been chemically... (Review)
Review
Pyoverdines are siderophores produced by fluorescent Pseudomonads to acquire iron. At least 60 different pyoverdines produced by diverse strains have been chemically characterized. They all consist of a dihydroquinoline-type chromophore linked to a peptide. These peptides are of various lengths and the sequences are strain specific. Pyoverdine biosynthesis in Pseudomonas aeruginosa and fluorescent Pseudomonads is a complex process involving at least 12 different proteins, starting in the cytoplasm and ending in the periplasm. The cellular localization of pyoverdine precursors was recently shown to be consistent with their biosynthetic enzymes. In the cytoplasm, pyoverdine appears to be assembled at the inner membrane and particularly at the old cell pole of the bacterium. Mature pyoverdine is uniformly distributed throughout the periplasm, like the periplasmic enzyme PvdQ. Secretion of pyoverdine involves a recently identified ATP-dependent efflux pump, PvdRT-OpmQ. This efflux system does not only secrete newly synthesized pyoverdine but also pyoverdine that already transported iron into the bacterial periplasm and any pyoverdine-metal complex other than ferri-pyoverdine present in the periplasm. This review considers how these new insights into pyoverdine biosynthesis and secretion contribute to our understanding of the role of pyoverdine in iron and metal homeostasis in fluorescent Pseudomonads.
Topics: Biological Transport; Cytoplasm; Homeostasis; Iron; Metals; Oligopeptides; Periplasm; Pseudomonas aeruginosa
PubMed: 23126435
DOI: 10.1111/1462-2920.12013 -
Trends in Microbiology Oct 2006Filamentous, heterocyst-forming cyanobacteria are multicellular organisms in which individual cells exchange nutrients and, presumably, regulatory molecules. Unknown...
Filamentous, heterocyst-forming cyanobacteria are multicellular organisms in which individual cells exchange nutrients and, presumably, regulatory molecules. Unknown mechanisms underlie this exchange. Classical electron microscopy shows that filamentous cyanobacteria bear a Gram-negative cell wall comprising a peptidoglycan layer and an outer membrane that are external to the cytoplasmic membrane, and that the outer membrane appears to be continuous along the filament of cells. This implies that the periplasmic space between the cytoplasmic and outer membranes might also be continuous. We propose that a continuous periplasm could constitute a communication conduit for the transfer of compounds, which is essential for the performance of these bacteria as multicellular organisms.
Topics: Cell Membrane; Cell Wall; Cyanobacteria; Periplasm
PubMed: 16934472
DOI: 10.1016/j.tim.2006.08.007