At conditions of low iron availability, Neisseria meningitidis produces a family of FrpC-like, type I-secreted RTX proteins of unknown role in meningococcal lifestyle. It is shown here that iron starvation also induces production of FrpD, the other protein expressed from a gene located immediately upstream of the frpC gene in a predicted iron-regulated frpDC operon. We found that FrpD is highly conserved in a set of meningococcal strains representative of all serogroups and does not exhibit any similarity to known sequences of other organisms. Subcellular localization and [3H]palmitic acid labeling in Escherichia coli revealed that FrpD is synthesized with a type II signal peptide for export across the cytoplasmic membrane and is, upon processing to a lipoprotein, sorted to the outer bacterial membrane. Furthermore, the biological function of FrpD appears to be linked to that of the RTX protein FrpC, because FrpD was found to bind the amino-proximal portion of FrpC (first 300 residues) with very high affinity (apparent Kd approximately 0.2 nM). These results suggest that FrpD represents an rtx loci-encoded accessory lipoprotein that could be involved in anchoring of the secreted RTX protein to the outer bacterial membrane.

• MeSH
• bakteriální proteiny chemie   genetika   metabolismus   MeSH
• konzervovaná sekvence MeSH
• lipoproteiny genetika   metabolismus   MeSH
• membránové proteiny chemie   genetika   metabolismus   MeSH
• molekulární sekvence - údaje MeSH
• Neisseria meningitidis genetika   metabolismus   MeSH
• proteiny vnější bakteriální membrány genetika   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• terciární struktura proteinů MeSH
• vápník farmakologie   MeSH
• vazba proteinů MeSH
• železo metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• frpC protein, Neisseria meningitidis MeSH Prohlížeč
• lipoproteiny MeSH
• membránové proteiny MeSH
• proteiny vnější bakteriální membrány MeSH
• vápník MeSH
• železo MeSH

Pseudomonas aeruginosa is one of the pathogenic bacteria which utilize binding of the host plasminogen (Plg) to promote their invasion throughout the host tissues. In the present study, we confirmed that P. aeruginosa exhibits binding affinity for human plasminogen. Furthermore, we showed that the protein detected on the cell wall of P. aeruginosa and binding human plasminogen is an enolase-like protein. The hypothesis that alpha-enolase, a cytoplasmatic glycolytic enzyme, resides also on the cell surface of the bacterium was supported by electron microscopy analysis. The plasminogen-binding activity of bacterial cell wall outer membrane enolase-like protein was examined by immunoblotting assay.

• MeSH
• fosfopyruváthydratasa metabolismus   MeSH
• imunoblotting MeSH
• imunoelektronová mikroskopie MeSH
• lidé MeSH
• plazminogen metabolismus   MeSH
• proteiny vnější bakteriální membrány metabolismus   MeSH
• Pseudomonas aeruginosa enzymologie   metabolismus   MeSH
• vazba proteinů MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fosfopyruváthydratasa MeSH
• plazminogen MeSH
• proteiny vnější bakteriální membrány MeSH

The periplasmic chaperone SurA plays a key role in outer membrane protein (OMP) biogenesis. E. coli SurA comprises a core domain and two peptidylprolyl isomerase domains (P1 and P2), but its mechanisms of client binding and chaperone function have remained unclear. Here, we use chemical cross-linking, hydrogen-deuterium exchange mass spectrometry, single-molecule FRET and molecular dynamics simulations to map the client binding site(s) on SurA and interrogate the role of conformational dynamics in OMP recognition. We demonstrate that SurA samples an array of conformations in solution in which P2 primarily lies closer to the core/P1 domains than suggested in the SurA crystal structure. OMP binding sites are located primarily in the core domain, and OMP binding results in conformational changes between the core/P1 domains. Together, the results suggest that unfolded OMP substrates bind in a cradle formed between the SurA domains, with structural flexibility between domains assisting OMP recognition, binding and release.

• MeSH
• Escherichia coli metabolismus   MeSH
• hmotnostní spektrometrie MeSH
• molekulární chaperony genetika   metabolismus   MeSH
• peptidylprolylisomerasa genetika   metabolismus   MeSH
• proteiny vnější bakteriální membrány genetika   metabolismus   MeSH
• proteiny z Escherichia coli genetika   metabolismus   MeSH
• transportní proteiny genetika   metabolismus   MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• molekulární chaperony MeSH
• peptidylprolylisomerasa MeSH
• proteiny vnější bakteriální membrány MeSH
• proteiny z Escherichia coli MeSH
• SurA protein, E coli MeSH Prohlížeč
• transportní proteiny MeSH

The iron-regulated FrpD protein is a unique lipoprotein embedded into the outer membrane of the Gram-negative bacterium Neisseria meningitidis. The biological function of FrpD remains unknown but might consist in anchoring to the bacterial cell surface the Type I-secreted FrpC protein, which belongs to a Repeat in ToXins (RTX) protein family and binds FrpD with very high affinity (K(d) = 0.2 nM). Here, we report the backbone (1)H, (13)C, and (15)N chemical shift assignments for the FrpD(43-271) protein that allow us to characterize the intimate interaction between FrpD and the N-terminal domain of FrpC.

• MeSH
• lipoproteiny chemie   MeSH
• Neisseria meningitidis metabolismus   MeSH
• nukleární magnetická rezonance biomolekulární * MeSH
• proteiny vnější bakteriální membrány chemie   MeSH
• sekvence aminokyselin MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• lipoproteiny MeSH
• proteiny vnější bakteriální membrány MeSH

To examine evidence of positive antibodies against immunogenic proteins of Anaplasma phagocytophilum in patients with other tick-borne infections and to diagnose possible co-infections, 412 serum specimens were tested by immunoblotting using three specific Anaplasma antigens: surface proteins p44 and Asp62 and outer membrane protein A (OmpA). In total, 284 serum samples from children with Lyme borreliosis and 12 serum samples from children with tick-borne encephalitis were tested. Sera from patients with viral aseptic meningitis (n = 47) and from blood donors (n = 69) were used as controls. Among all serum specimens from patients with tick-borne infections submitted for this study, six samples (2·0%) showed positive IgM reactions and seven samples (2·4%) were IgG positive for A. phagocytophilum by immunoblot. Borderline reactivity was found in 30 samples (10·14%) for IgM and 36 samples (12·2%) for IgG. The difference between patients and blood donors was statistically significant for IgM (P = 0·006) and for IgG (P = 0·0007) antibodies. A statistically significant result was obtained for IgG (P = 0·02) but not for IgM between patients and children with aseptic meningitis. Immunoblot using three specific antigens provides novel information about the positivity of antibodies to A. phagocytophilum in children with other tick-borne infections. Taking into account clinical and laboratory findings of children despite antibody positivity, no case of human granulocytic anaplasmosis was demonstrated.

• Klíčová slova
• Anaplasma phagocytophilum, Lyme borreliosis, human granulocytic anaplasmosis, major surface proteins, outer membrane protein A, tick-borne encephalitis,
• MeSH
• Anaplasma phagocytophilum imunologie   MeSH
• anaplasmóza diagnóza   imunologie   mikrobiologie   MeSH
• bakteriální proteiny imunologie   MeSH
• dítě MeSH
• klíšťová encefalitida imunologie   mikrobiologie   MeSH
• koinfekce diagnóza   imunologie   mikrobiologie   MeSH
• lidé MeSH
• lymeská nemoc imunologie   mikrobiologie   MeSH
• membránové proteiny imunologie   MeSH
• mladiství MeSH
• předškolní dítě MeSH
• proteiny vnější bakteriální membrány imunologie   MeSH
• protilátky bakteriální krev   MeSH
• western blotting MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• bakteriální proteiny MeSH
• membránové proteiny MeSH
• proteiny vnější bakteriální membrány MeSH
• protilátky bakteriální MeSH

Francisella tularensis is a highly infectious Gram-negative coccobacillus which causes the disease tularemia. The potential for its misuse as a biological weapon has led disease control and prevention centers to classify this bacterium as a category A agent. Bacterial outer membrane vesicles (OMVs) are spherical particles 20-250 nm in size produced by all Gram-negative bacteria and constitute one of the major secretory pathways. Bacteria use them in interacting with both other bacterial cells and eukaryotic (host) cells. OMVs of Francisella contain number of its so far described virulence factors and immunomodulatory proteins. Their role in host-pathogen interactions can therefore be presumed, and the possibility exists also for their potential use in a subunit vaccine. Moreover, Francisella microbes produce both usual spherical and unusual tubular OMVs. Because OMVs emerge from the outermost surface of the bacterial cell, we focused on the secretion of OMVs in several mutant Francisella strains with disrupted surface structures (namely the O-antigen). O-antigen in Francisella is not only the structural component of LPS but also forms another important virulence factor: the O-antigen polysaccharide capsule. Mutant strain phenotypes were evaluated by growth curves, vesiculation rates, their sensitivity to the complement contained in serum, and proliferation inside murine bone marrow macrophages. Morphologies of both OMVs and the bacteria were visualized by electron microscopy. The O-antigen mutant strains were considerably attenuated in serum resistance and intracellular proliferation. All the strains showed lower ability to form the tubular OMVs. Some strains formed tubular protrusions from their outer membrane but their stability was weak. Some hypervesiculating strains were revealed that will serve as source of OMVs for further studies of their protective potential. Our results suggest the presence of LPS and the O-antigen capsule on the surface of Francisella to be critical not only for its virulence but also for the exceptional tubular shape of its OMVs.

• Klíčová slova
• Capsule, Francisella tularensis, Lipopolysaccharide, Nanotubes, O-antigen, Outer membrane vesicles,
• MeSH
• Francisella tularensis * genetika   MeSH
• gramnegativní bakterie MeSH
• lipopolysacharidy chemie   MeSH
• myši MeSH
• O-antigeny MeSH
• proteiny vnější bakteriální membrány genetika   metabolismus   MeSH
• tularemie * mikrobiologie   prevence a kontrola   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• lipopolysacharidy MeSH
• O-antigeny MeSH
• proteiny vnější bakteriální membrány MeSH

Fe-regulated protein D (FrpD) is a Neisseria meningitidis outer membrane lipoprotein that may be involved in the anchoring of the secreted repeat in toxins (RTX) protein FrpC to the outer bacterial membrane. However, the function and biological roles of the FrpD and FrpC proteins remain unknown. Native and selenomethionine-substituted variants of recombinant FrpD43-271 protein were crystallized using the sitting-drop vapour-diffusion method. Diffraction data were collected to a resolution of 2.25 A for native FrpD43-271 protein and to a resolution of 2.00 A for selenomethionine-substituted FrpD43-271 (SeMet FrpD43-271) protein. The crystals of native FrpD43-271 protein belonged to the hexagonal space group P6(2) or P6(4), while the crystals of SeMet FrpD43-271 protein belonged to the primitive orthorhombic space group P2(1)2(1)2(1).

• MeSH
• krystalizace MeSH
• krystalografie rentgenová MeSH
• lipoproteiny chemie   MeSH
• Neisseria meningitidis chemie   MeSH
• proteiny vnější bakteriální membrány chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• lipoproteiny MeSH
• proteiny vnější bakteriální membrány MeSH

In recent years, considerable progress has been made in topologically and functionally characterizing integral outer membrane proteins (OMPs) of Treponema pallidum subspecies pallidum, the syphilis spirochete, and identifying its surface-exposed β-barrel domains. Extracellular loops in OMPs of Gram-negative bacteria are known to be highly variable. We examined the sequence diversity of β-barrel-encoding regions of tprC, tprD, and bamA in 31 specimens from Cali, Colombia; San Francisco, California; and the Czech Republic and compared them to allelic variants in the 41 reference genomes in the NCBI database. To establish a phylogenetic framework, we used T. pallidum 0548 (tp0548) genotyping and tp0558 sequences to assign strains to the Nichols or SS14 clades. We found that (i) β-barrels in clinical strains could be grouped according to allelic variants in T. pallidum subsp. pallidum reference genomes; (ii) for all three OMP loci, clinical strains within the Nichols or SS14 clades often harbored β-barrel variants that differed from the Nichols and SS14 reference strains; and (iii) OMP variable regions often reside in predicted extracellular loops containing B-cell epitopes. On the basis of structural models, nonconservative amino acid substitutions in predicted transmembrane β-strands of T. pallidum repeat C (TprC) and TprD2 could give rise to functional differences in their porin channels. OMP profiles of some clinical strains were mosaics of different reference strains and did not correlate with results from enhanced molecular typing. Our observations suggest that human host selection pressures drive T. pallidum subsp. pallidum OMP diversity and that genetic exchange contributes to the evolutionary biology of T. pallidum subsp. pallidum They also set the stage for topology-based analysis of antibody responses to OMPs and help frame strategies for syphilis vaccine development.IMPORTANCE Despite recent progress characterizing outer membrane proteins (OMPs) of Treponema pallidum, little is known about how their surface-exposed, β-barrel-forming domains vary among strains circulating within high-risk populations. In this study, sequences for the β-barrel-encoding regions of three OMP loci, tprC, tprD, and bamA, in T. pallidum subsp. pallidum isolates from a large number of patient specimens from geographically disparate sites were examined. Structural models predict that sequence variation within β-barrel domains occurs predominantly within predicted extracellular loops. Amino acid substitutions in predicted transmembrane strands that could potentially affect porin channel function were also noted. Our findings suggest that selection pressures exerted within human populations drive T. pallidum subsp. pallidum OMP diversity and that recombination at OMP loci contributes to the evolutionary biology of syphilis spirochetes. These results also set the stage for topology-based analysis of antibody responses that promote clearance of T. pallidum subsp. pallidum and frame strategies for vaccine development based upon conserved OMP extracellular loops.

• Klíčová slova
• Treponema pallidum, molecular subtyping, outer membrane proteins, spirochetes, syphilis,
• MeSH
• fylogeneze MeSH
• genetická variace MeSH
• lidé MeSH
• molekulární evoluce * MeSH
• molekulární sekvence - údaje MeSH
• proteinové domény MeSH
• proteiny vnější bakteriální membrány chemie   genetika   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční seřazení MeSH
• Spirochaetales klasifikace   genetika   růst a vývoj   izolace a purifikace   MeSH
• syfilis mikrobiologie   MeSH
• Treponema pallidum klasifikace   genetika   růst a vývoj   izolace a purifikace   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• proteiny vnější bakteriální membrány MeSH

The aim of this work was isolation and purification of the major immunodominant protein, Outer surface protein C (OspC) of three members of the species group Borrelia burgdorferi, the causative agent of Lyme disease. Our aim was to obtain this protein in a quantity and purity sufficient for immunization of experimental animals. For optimalization of protein purification's yield we used immobilized metal ion affinity chromatography (IMAC) under different conditions. The greatest efficiency was achieved by using of HiTrap Chelating Column under native conditions.

• MeSH
• antigeny bakteriální biosyntéza   izolace a purifikace   MeSH
• DNA vakcíny biosyntéza   izolace a purifikace   MeSH
• Escherichia coli MeSH
• genetické vektory MeSH
• proteiny vnější bakteriální membrány biosyntéza   izolace a purifikace   MeSH
• vakcína proti lymeské nemoci biosyntéza   izolace a purifikace   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antigeny bakteriální MeSH
• DNA vakcíny MeSH
• OspC protein MeSH Prohlížeč
• proteiny vnější bakteriální membrány MeSH
• vakcína proti lymeské nemoci MeSH

Hydrophobicity and profiles of outer membrane proteins of Shigella dysenteriae type 1 after treatment with subinhibitory concentrations (1/2 or 1/4 of the MIC) of aminoglycosides were studied. The antimicrobial activity of the antibiotics tested was 3.12 mg/L (amikacin, tobramycin) and 6.25 mg/L (gentamicin). The hydrophobicity of the cell surface of S. dysenteriae type 1 was decreased after exposure to all aminoglycosides at a concentration of 1/2 of the MICs; 1/4 of the MICs of the antibiotics did not affect bacterial aggregation in the presence of ammonium sulfate. SDS-polyacrylamide gel electrophoresis showed that the profiles of outer membrane proteins of the strain treated with aminoglycosides at both subinhibitory concentrations were not changed as compared to the control.

• MeSH
• amikacin aplikace a dávkování   farmakologie   MeSH
• antibakteriální látky farmakologie   MeSH
• buněčná stěna chemie   účinky léků   MeSH
• chemické jevy MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• fyzikální chemie MeSH
• gentamiciny aplikace a dávkování   farmakologie   MeSH
• proteiny vnější bakteriální membrány chemie   MeSH
• Shigella dysenteriae účinky léků   metabolismus   MeSH
• tobramycin aplikace a dávkování   farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• srovnávací studie MeSH
• Názvy látek
• amikacin MeSH
• antibakteriální látky MeSH
• gentamiciny MeSH
• proteiny vnější bakteriální membrány MeSH
• tobramycin MeSH