| 1. |
- Janulczyk, Robert, et al.
(författare)
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Improved pattern for genome-based screening identifies novel cell wall-attached proteins in gram-positive bacteria
- 2001
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Ingår i: Infection and Immunity. - 1098-5522. ; 69:6, s. 4019-4026
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Tidskriftsartikel (refereegranskat)abstract
- With a large number of sequenced microbial genomes available, tools for identifying groups or classes of proteins have become increasingly important. Here we present an improved pattern for the identification of cell wall-attached proteins (CWPs), a group of proteins with diverse and important functions in gram-positive bacteria. This tripartite pattern is based on analysis of 65 previously described cell wall-attached proteins and takes into account the three principal requirements for cell wall sorting; a sortase target region (LPXTGX), a membrane-spanning region, and a charged stop-transfer tail. In five different genomes of gram-positive bacteria, the tripartite pattern identified a total of 35 putative CWPs, 19 of which were novel. The specificity and sensitivity of the tripartite pattern are higher than those of the classical pattern, which is based solely on the sortase target region. Several putative CWPs with atypical sortase target regions were identified. In the complete genome of the important human pathogen Streptococcus pyogenes, the tripartite pattern identified 14 putative CWPs. Seven of the putative S. pyogenes proteins were novel, and two of these were a 5' nucleotidase and a pullulanase. This study represents the first whole-genome screening for CWPs, and we conclude that the tripartite pattern is highly suitable for this purpose. Identification of CWPs using this pattern offers important possibilities in the study of the pathogenesis and physiology of gram-positive bacteria.
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| 2. |
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| 3. |
- Janulczyk, Robert
(författare)
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Studies on surface proteins of Gram-positive bacteria
- 2001
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Despite the availability of antibiotics and modern health care, infectious diseases continue to cause great suffering and costs. Bacterial surface proteins are important for the interaction between host and pathogen. This thesis gives an overview of the importance of bacterial surface proteins in Gram-positive bacteria, particularly Streptococcus pneumoniae and Streptococcus pyogenes. A novel contribution is the identification and characterization of a metal transport system in S. pyogenes, with broad specificity for trace metals. We have showed that this transporter, MtsABC, mediates bacterial acquisition of iron, zinc, and manganese. A MtsABC-deficient strain had attenuated virulence, and was highly susceptible to oxidative stress. We also observed reduced functionality of a manganese-dependent superoxide dismutase. Another contribution is the identification of a factor H-binding inhibitor of complement (Hic) in S. pneumoniae. This surface protein is encoded by the pspC locus, but homology between Hic and PspC is limited to the N-terminal part of the proteins. Hic binds fH with high affinity, and without interfering with the complement inhibitory effect of fH. FH is a major complement regulatory protein, and recruitment of fH to the bacterial surface may impair the activation of the alternative pathway. A bioinformatic work is also presented, where we develop and refine a pattern for computerized identification of cell wall-attached proteins (CWPs) in genomes from Gram-positive bacteria. Application of the pattern resulted in identification of 35 genes encoding putative CWPs, 19 of which were previously unknown.
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| 4. |
- Jarva, Hanna, et al.
(författare)
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Streptococcus pneumoniae evades complement attack and opsonophagocytosis by expressing the pspC locus-encoded Hic protein that binds to short consensus repeats 8-11 of factor H
- 2002
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Ingår i: Journal of Immunology. - 1550-6606. ; 168:4, s. 1886-1894
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Tidskriftsartikel (refereegranskat)abstract
- Streptococcus pneumoniae is an important cause of upper and lower respiratory tract infections, meningitis, peritonitis, bacterial arthritis, and sepsis. Here we have studied a novel immune evasion mechanism of serotype 3 pneumococci, which are particularly resistant to phagocytosis. On their surfaces the bacteria express the factor H-binding inhibitor of complement (Hie), a protein of the pneumococcal surface protein C family. Using radioligand binding, microtiter plate assays, surface plasmon resonance analysis, and recombinant constructs of factor H, we located the binding site of Hie to short consensus repeats (SCRs) 8-11 in the middle part of factor H. This represents a novel microbial interaction region on factor H. The only other ligand known so far for SCRs 8-11 of factor H is C-reactive protein (CRP), an acute phase protein that binds to the pneumococcal C-polysaccharide. The binding sites of Hie and CRP within the SCR8-11 region were different, however, because CRP did not inhibit the binding of Hie and required calcium for binding. Binding of factor H to Hic-expressing pneumococci promoted factor I-mediated cleavage of C3b and restricted phagocytosis of pneumococci. Thus, virulent pneumococci avoid complement attack and opsonophagocytosis by recruiting functionally active factor H with the Hie surface protein. Hie binds to a previously unrecognized microbial interaction site in the middle part of factor H.
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| 5. |
- Linder, Adam, et al.
(författare)
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Human antibody response towards the pneumococcal surface proteins PspA and PspC during invasive pneumococcal infection.
- 2007
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Ingår i: Vaccine. - : Elsevier BV. - 1873-2518 .- 0264-410X. ; 25:2, s. 341-345
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Tidskriftsartikel (refereegranskat)abstract
- gG antibodies against pneumococcal surface protein A, family 1 (PspAl) and family 2 (PspA2), protein C (PspC), and protein Hic were investigated in 41 patients with invasive pneumococcal disease. Pre-existing antibody levels against the four pneumococcal proteins were not significantly different from those found in 40 patients with non-pneumococcal bacteremia or 80 healthy controls. However, during convalescense a strong immune response developed especially against PspA, and there was a high degree of cross-reactivity between PspA-and PspC-antibodies. Our findings on immunogenicity and cross-reactivity suggest that in a future pneumococcal protein based vaccine, only a limited number of proteins could be sufficient.
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| 6. |
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| 7. |
- Murphy, Elizabeth, et al.
(författare)
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Identification of pili on the surface of Finegoldia magna - A Gram-positive anaerobic cocci.
- 2014
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Ingår i: Anaerobe. - : Elsevier BV. - 1095-8274 .- 1075-9964. ; 27:Mar 29, s. 40-49
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Tidskriftsartikel (refereegranskat)abstract
- Pili have only been discovered in the major Gram-positive pathogens in the past decade and they have been found to play an important role in colonisation and virulence. Pili have been shown to have many important functions including attachment to host tissues, mediating bacterial aggregation, biofilm formation and binding to proteins in the extracellular matrix. In this study, sortase-dependent pili have been found to be expressed on the surface of Finegoldia magna ALB8. F. magna is a Gram-positive anaerobic coccus that, primarily, is a commensal of the skin and mucous membranes, but has also been isolated from various clinical infection sites and is associated with soft-tissue abscesses, wound infections and bone and prosthetic joint infections. In this study, F. magna ALB8 was found to harbour three sortases at the pilus locus, two of which bear high similarity to class C sortases in Streptococcus pneumoniae. Two putative sortase-dependent pili proteins were found in the locus, with one being identified as the major pilus subunit, Fmp1 (F. magna pilus subunit 1), due to its high similarity to other major pilus proteins in prominent Gram-positive pathogens. The presence of sortase-dependent pili was confirmed experimentally through recombinant production of Fmp1 and production of antiserum. The Fmp1 antiserum was used in Western blot to show the presence of a high molecular weight protein ladder, characteristic of the presence of pili, in trypsin released cell wall surface proteins from F. magna. The presence of sortase-dependent pili was visually confirmed by transmission electron microscopy, which showed the binding of gold labelled anti-Fmp1 to individual pilus proteins along the pilus. Furthermore, pili could also be found to bind and interact with keratinocytes in the epidermal layer of human skin, suggesting an adhesive role for pili on F. magna. Our work represents the first description of pilus structures in F. magna. This discovery further elucidates F. magna physiology and allows for additional analysis of host-bacterial interactions in future studies.
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| 8. |
- Ricci, Susanna, et al.
(författare)
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The factor H-binding fragment of PspC as a vaccine antigen for the induction of protective humoral immunity against experimental pneumococcal sepsis
- 2011
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Ingår i: Vaccine. - : Elsevier BV. - 1873-2518 .- 0264-410X. ; 29:46, s. 8241-8249
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Tidskriftsartikel (refereegranskat)abstract
- Pneumococcal surface protein C (PspC) is a major virulence factor of Streptococcus pneumoniae and interferes with complement activity by binding complement factor H (fH). In this study, protection against experimental sepsis caused by pneumococci carrying different PspC variants was evaluated by immunisation with the fH-binding fragment of PspC. The mechanisms of protection mediated by antibodies to PspC were also studied. Mice were immunised with a PspC fragment (PspC(39-261)) from the type 3 strain HB565 and infected intravenously with either strain HB565 (homologous challenge), or strains D39 and TiGR4 (heterologous challenge). Immunisation with PspC(39-261) elicited high titers (>300,000) of PspC-specific serum IgG and conferred protection from challenge with HB565. In contrast, cross-protection was either limited or absent in vaccinated animals infected with D39 and TIGR4, respectively. To correlate protection with reactivity and function of PspC antibodies, pooled sera from vaccinated mice were tested in IgG binding and complement deposition experiments. IgG antibodies efficiently bound to HB565, while binding was lower with D39 and absent with TIGR4. In the presence of mouse post-immune sera, C3 deposition was increased onto HB565, while no effect was observed with 039 and TIGR4. Antibody cross-reactivity and complement deposition progressively declined with reduced amino acid identity between PspC variants. Antibodies to PspC were also found to interfere with binding to HB565. Finally, in vitro and ex vivo phagocytosis assays demonstrated that PspC-specific antibodies promoted opsonophagocytic killing of bacteria. (C) 2011 Elsevier Ltd. All rights reserved.
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| 9. |
- Ricci, Susanna, et al.
(författare)
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The regulator PerR is involved in oxidative stress response and iron homeostasis and is necessary for full virulence of Streptococcus pyogenes.
- 2002
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Ingår i: Infection and Immunity. - 1098-5522. ; 70:9, s. 4968-4976
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Tidskriftsartikel (refereegranskat)abstract
- Ferric uptake regulator (Fur) and Fur-like proteins form an important family of transcriptional regulators in many bacterial species. In this work we have characterized a Fur-like protein, the peroxide regulator PerR, in an M1 serotype of Streptococcus pyogenes. To determine the role of PerR in S. pyogenes, we inactivated the gene by allelic replacement. PerR-deficient bacteria showed 48% reduction of (55)Fe incorporation from the culture medium. Transcriptional analysis revealed that mtsA, encoding a metal-binding protein of an ABC transporter in S. pyogenes, was transcribed at lower levels than were wild-type cells. Although total iron accumulation was reduced, the growth of the mutant strain was not significantly hampered. The mutant showed hyperresistance to hydrogen peroxide, and this response was induced in wild-type cells by growth in aerobiosis, suggesting that PerR acts as an oxidative stress-responsive repressor. PerR may also participate in the response to superoxide stress, as the perR mutant was more sensitive to the superoxide anion and had a reduced transcription of sodA, which encodes the sole superoxide dismutase of S. pyogenes. Complementation of the mutation with a functional perR gene restored (55)Fe incorporation, response to peroxide stress, and transcription of both mtsA and sodA to levels comparable to those of wild-type bacteria. Finally, the perR mutant was attenuated in virulence in a murine air sac model of infection (P < 0.05). These results demonstrate that PerR is involved in the regulation of iron homeostasis and oxidative stress responses and that it contributes to the virulence of S. pyogenes.
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| 10. |
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