Expression, purification and metal utilization of recombinant SodA from Borrelia burgdorferi

Publication date: Available online 1 July 2019Source: Protein Expression and PurificationAuthor(s): G. Brown, A.H. Broxham, S.E. Cherrington, D.C. Thomas, A. Dyer, L. Stejskal, R.J. BinghamAbstractBorrelia are microaerophilic spirochetes capable of causing multisystemic diseases such as Lyme disease and Relapsing Fever. The ubiquitous Fe/Mn-dependent superoxide dismutase (SOD) provides essential protection from oxidative damage by the superoxide anion. Borrelia possess a single SOD enzyme - SodA that is essential for virulence, providing protection against host-derived reactive oxygen species (ROS). Here we present a method for recombinant expression and purification of Borrelia burgdorferi SodA in E. coli. Metal exchange or insertion into the Fe/Mn-SOD is inhibited in the folded state. We therefore present a method whereby the recombinant Borrelia SodA binds to Mn under denaturing conditions and is subsequently refolded by a reduction in denaturant. SodA purified by metal affinity chromatography and size exclusion chromatography reveals a single band on SDS-PAGE. Protein folding is confirmed by circular dichroism. A coupled enzyme assay demonstrates SOD activity in the presence of Mn, but not Fe. The apparent molecular weight determined by size exclusion corresponds to a dimer of SodA; a homology model of dimeric SodA is presented revealing a surface Cys distal to the dimer interface. The method presented of acquiring a target metal under denaturing conditions may be applica...
Source: Protein Expression and Purification - Category: Biochemistry Source Type: research

Related Links:

In this study, the proteomic profile of the surface-associated proteins from M. bovis BCG Moreau was compared to the BCG Pasteur reference strain. The methodology used was 2DE gel electrophoresis combined with mass spectrometry techniques (MALDI-TOF/TOF), leading to the identification of 115 proteins. Of these, 24 proteins showed differential expression between the two BCG strains. Furthermore, 27 proteins previously described as displaying moonlighting function were identified, 8 of these proteins showed variation in abundance comparing BCG Moreau to Pasteur and 2 of them presented two different domain hits. Moonlighting ...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
Charlotte Mason1, Xiaoyan Liu2, Spoorthy Prabhudeva1 and Zhiming Ouyang1* 1Department of Molecular Medicine, University of South Florida, Tampa, FL, United States 2Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX, United States BosR, a Fur family member, is essential for the pathogenesis of the Lyme disease pathogen, Borrelia burgdorferi. Unlike typical Fur proteins in which DNA binding represses gene expression, binding of BosR to the rpoS promoter directly activates rpoS transcription in B. burgdorferi. However, virtually nothing is known concerning potential structural fea...
Source: Frontiers in cellular and infection microbiology - Category: Microbiology Source Type: research
Tania Rahman1,2†‡, Andrew S. Brown2‡, Elizabeth L. Hartland1‡, Ian R. van Driel2* and Ka Yee Fung1†‡ 1Department of Microbiology and Immunology, University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, VIC, Australia 2Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Melbourne, VIC, Australia We have examined the influence of depleting plasmacytoid dendritic cells (pDC) in mice on the immune response to the gut pathogen Citrobacter rodentium, an organism tha...
Source: Frontiers in Immunology - Category: Allergy & Immunology Source Type: research
CONCLUSIONS: Immunization with native antigens might not be sufficient to obtain complete protection to infection. Nonetheless we showed that non-infectious B. burgdorferi can be an effective carrier to deliver and elicit a specific host response to T. pallidum antigens to assess the efficacy of syphilis vaccine candidates. PMID: 30797635 [PubMed - as supplied by publisher]
Source: Vaccine - Category: Allergy & Immunology Authors: Tags: Vaccine Source Type: research
In this study, bird species migrating through Bulgaria were investigated as carriers of zoonotic pathogens. In total, 706 birds belonging to 46 species were checked for the presence of various bacterial pathogens (Campylobacter, Yersin ia, Salmonella, Listeria, Escherichia coli, Staphylococcus aureus, Francisella tularensis, Coxiella burnetii, Borrelia burgdorferi, and Brucella spp.). From 673 birds we investigated fecal samples, from the remaining 33, blood samples. We detected Campylobacter 16S rDNA gene in 1.3% of birds, but no ne were of pathogenic Campylobacter jejuni and Campylobacter coli species. Escherichia coli 1...
Source: Canadian Journal of Microbiology - Category: Microbiology Authors: Source Type: research
Answer:Borreliaspp., relapsing fever group. Molecular studies are needed to definitively identify the causative agent. However, knowing the exposure history of the patient also helps narrow the differential. In this case, we later found out that the patient is from the northwestern part of North America, and therefore this is most likelyBorrelia hermsii,one of the tick-borne relapsing feverBorreliaspecies.The other tick-borneBorreliain North America areB. parkeriiandB. turicatae.These species are transmitted through the bite of a soft-bodied tick in the genusOrnithodoros,and are usually associated with exposure to'rustic's...
Source: Creepy Dreadful Wonderful Parasites - Category: Parasitology Source Type: blogs
In this study, the chimeric proteins (A/C-2, A/C-4 and A/C-7.1) consisting of B-cell epitopes of outer surface proteins OspA and OspC from Borrelia genospecies prevalent in Eastern Slovakia, were designed, over-expressed in E. coli, and used to detect specific anti-Borrelia antibodies in serologically characterized sera from patients with Lyme-like symptoms to evaluate their diagnostic potential. RESULTS: Results showed that chimeras vary in their immuno-reactivity when tested with human sera. Compared with the results obtained from a two-tier test, the application of recombinant multi-epitope chimeric proteins as dia...
Source: Annals of Agricultural and Environmental Medicine : AAEM - Category: Environmental Health Authors: Tags: Ann Agric Environ Med Source Type: research
ConclusionsThere was a very high prevalence of D. immitis, hookworm and external parasites in Samoan dogs, and prophylactic medication is rarely administered. This is the first report confirming A. platys in Samoa and the South Pacific islands. The public health implications of poor management of the dogs should be considered and investigated further.
Source: Australian Veterinary Journal - Category: Veterinary Research Authors: Tags: SMALL ANIMALS Source Type: research
Conclusions: The results of detection and attribution studies can inform evidence-based risk management to reduce current, and plan for future, changes in health risks associated with climate change. Gaining a better understanding of the size, timing, and distribution of the climate change burden of disease and injury requires reliable long-term data sets, more knowledge about the factors that confound and modify the effects of climate on health, and refinement of analytic techniques for detection and attribution. At the same time, significant advances are possible in the absence of complete data and statistical certainty:...
Source: EHP Research - Category: Environmental Health Authors: Tags: Commentary Source Type: research
Publication date: Available online 28 January 2017 Source:Best Practice & Research Clinical Haematology Author(s): Maurilio Ponzoni, Andrés JM. Ferreri In the last decades some bacteria have been associated with a various extent with marginal zone (extra nodal, nodal and splenic types) lymphomas are frequently associated with chronic infections, with important clinical, molecular, biological, and therapeutic implications. The well-known correlation between Helicobacter pylori and gastric MALT-lymphoma, the recently reported links between Chlamydia psittaci and ocular adnexal MALT-lymphoma and Borrelia burgdorfe...
Source: Best Practice and Research Clinical Haematology - Category: Hematology Source Type: research
More News: Biochemistry | Borrelia | Gastroenteritis | Lyme Disease