1,720,991 research outputs found
Structural vaccinology for melioidosis vaccine design and immunodiagnostics
Full text linkPurpose of Review
Spurred by the successful application of structural vaccinology to other challenging bacterial and viral pathogens, we review the possibility of exploiting 3D structure computational-based recombinant antigen engineering strategies for the development of a protective melioidosis vaccine.
Recent Findings
Structure-based epitope design approaches in the melioidosis field are preliminary and applied essentially by one research network. By combining Burkholderia pseudomallei antigen 3D structures and in silico epitope discovery methods, a panel of synthetic epitope peptides were designed and tested for their B and T cell stimulatory activities. Several peptides were found to be serodiagnostic for B. pseudomallei infection and two elicited bactericidal antibodies.
Summary
A significant amount of B. pseudomallei antigen structures, epitopes, and immunological data is available. Future challenges will be to test all available B. pseudomallei epitopes, focusing on combing multiple B/T cell epitopes onto a single scaffold to generate components, stimulating both arms of the immune system
Group B Streptococcus pullulanase crystal structures in the context of a novel strategy for vaccine development
No full textThe group B streptococcus type I pullulanase (SAP) is a class 13 glycoside hydrolase that is anchored to the bacterial cell surface via a conserved C-terminal anchoring motif and involved in {alpha}-glucan degradation. Recent in vitro functional studies have shown that SAP is immunogenic in humans and that anti-SAP sera derived from immunized animals impair both group A and group B streptococcus pullulanase activities, suggesting that in vivo immunization with this antigen could prevent streptococcal colonization. To further investigate the putative role of SAP in bacterial pathogenesis, we carried out functional studies and found that recombinant SAP binds to human cervical epithelial cells. Furthermore, with a view of using SAP as a vaccine candidate, we present high-resolution crystal structure analyses of an N-terminally truncated form of SAP lacking the carbohydrate binding module but containing the catalytic domain and displaying glycosidase hydrolase activity, both in its apo form and in complex with maltotetraose, at resolutions of 2.1 and 2.4 Å, respectively
Why is a protective antigen protective?
No full textOne of the major challenges in vaccine development is the identification of microbial components that give rise to a protective immune response. Over the last decade, genome and proteome-based methods have proven successful in discovering new vaccine candidates for many pathogens through the selection of secreted or surface-exposed protein antigens and their screening in proper biological assays. However, these approaches still require intensive research activities to single out, among the large number of secreted and surface exposed proteins those very few which are protective. The question of which structural properties render an antigen capable of eliciting the production of functional remains most challenging. In such scientific and methodological context, the EU-funded project BacAbs was set up in 2007 with the main objective of developing a knowledge-based protocol able to discern protective from non-protective protein antigens, based on their amino acid sequences and molecular properties. The successful BacAbs multidisciplinary approach is highlighted by the recent analysis of three antigens from two diverse pathogens. Here, we describe the work and results carried out so far by the BacAbs Consortium, and discuss its relevance with regards to accelerating antigen selection processes and state-of-the-art vaccine development
Backbone and side-chain (1)H, (15)N, (13)C assignment and secondary structure of BPSL1445 from Burkholderia pseudomallei
No full textBPSL1445 is a lipoprotein produced by the Gram-negative bacterium Burkholderia pseudomallei (B.pseudomallei), the etiological agent of melioidosis. Immunodetection assays against sera patients using protein microarray suggest BPSL1445 involvement in melioidosis. Herein we report backbone, side chain NMR assignment and secondary structure for the recombinant protein
Structure-function analysis of recombinant substrate protein 22 kDa (SP-22) - A mitochondrial 2-Cys peroxiredoxin organized as a decameric toroid
No full textBovine mitochondrial SP-22 is a member of the peroxiredoxin family of peroxidases. It belongs to the peroxiredoxin 2-Cys subgroup containing three cysteines at positions 47, 66, and 168. The cloning and overexpression in Escherichia coli of recombinant wild type SP-22 and its three cysteine mutants (C47S, C66S, and C168S) are reported. Purified His-tagged SP-22 was fully active with Cys-47 being confirmed as the catalytic residue. The enzyme forms a stable decameric toroid consisting of five basic dimeric units containing intermolecular disulfide bonds linking the catalytically active Cys-47 of one subunit and Cys-168 of the adjacent monomer. The disulfide bonds are not required for overall structural integrity. The toroidal units have average external and internal diameters of 15 and 7 nm, respectively, and can form stacks in a lateral arrangement of two or three rings. C47S had a pronounced tendency to stack in long tubular structures containing up to 60 rings. Further unusual structural features are the presence of radial spikes projecting from the external surface and ordered electron-dense material within the central cavity of the toroid
Bovine mitochondrial peroxiredoxin III forms a two-ring catenane
No full textSummaryA crystal structure is reported for the C168S mutant of a typical 2-Cys peroxiredoxin III (Prx III) from bovine mitochondria at a resolution of 3.3 Å. Prx III is present as a two-ring catenane comprising two interlocking dodecameric toroids that are assembled from basic dimeric units. Each ring has an external diameter of 150 Å and encompasses a central cavity that is 70 Å in width. The concatenated dodecamers are inclined at an angle of 55°, which provides a large contact surface between the rings. Dimer-dimer contacts involved in toroid formation are hydrophobic in nature, whereas the 12 areas of contact between interlocked rings arise from polar interactions. These two major modes of subunit interaction provide important insights into possible mechanisms of catenane formation
Crystal structure of an R-selective transaminase from Thermomyces stellatus
No full texthttps://www.wwpdb.org/pdb?id=pdb_00006xw
Boosting of post-exposure human T-cell and B-cell recall responses in vivo by Burkholderia pseudomallei-related proteins.
Full text linkBurkholderia pseudomallei is the causative agent of melioidosis, an infectious disease with high incidence and mortality in South East Asia and northern Australia. To date there is no protective vaccine and antibiotic treatment is prolonged and not always effective. Most people living in endemic areas have been exposed to the bacteria and have developed some immunity, which may have helped to prevent disease. Here, we used a humanized mouse model (hu-PBL-SCID), reconstituted with human peripheral blood mononuclear cells from seropositive donors, to illustrate the potential of three known antigens (FliC, OmpA and N-PilO2) for boosting both T-cell and B-cell immune responses. All three antigens boosted the production of specific antibodies in vivo, and increased the number of antibody and interferon-γ-secreting cells, and induced antibody affinity maturation. Moreover, antigen-specific antibodies isolated from either seropositive individuals or boosted mice, were found to enhance phagocytosis and oxidative burst activities from human polymorphonuclear cells. Our study demonstrates that FliC, OmpA and N-PilO2 can stimulate human memory T and B cells and highlight the potential of the hu-PBL-SCID system for screening and evaluation of novel protein antigens for inclusion in future vaccine trials against melioidosis
The Escherichia coli Lpt transenvelope protein complex for lipopolysaccharide export is assembled via conserved structurally homologous domains
Full text linkLipopolysaccharide is a major glycolipid component in the outer leaflet of the outer membrane (OM), a peculiar permeability barrier of Gram-negative bacteria that prevents many toxic compounds from entering the cell. Lipopolysaccharide transport (Lpt) across the periplasmic space and its assembly at the Escherichia coli cell surface are carried out by a transenvelope complex of seven essential Lpt proteins spanning the inner membrane (LptBCFG), the periplasm (LptA), and the OM (LptDE), which appears to operate as a unique machinery. LptC is an essential inner membrane-anchored protein with a large periplasm-protruding domain. LptC binds the inner membrane LptBFG ABC transporter and interacts with the periplasmic protein LptA. However, its role in lipopolysaccharide transport is unclear. Here we show that LptC lacking the transmembrane region is viable and can bind the LptBFG inner membrane complex; thus, the essential LptC functions are located in the periplasmic domain. In addition, we characterize two previously described inactive single mutations at two conserved glycines (G56V and G153R, respectively) of the LptC periplasmic domain, showing that neither mutant is able to assemble the transenvelope machinery. However, while LptCG56V failed to copurify any Lpt component, LptCG153R was able to interact with the inner membrane protein complex LptBFG. Overall, our data further support the model whereby the bridge connecting the inner and outer membranes would be based on the conserved structurally homologous jellyroll domain shared by five out of the seven Lpt components
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