Project description:Several two-component systems of Streptomyces coelicolor, a model organism to study antibiotic production in Streptomyces, affects the expression of the bfr (SCO2113) gene, which codifies for a bacterioferritin, a protein involved in iron storage. The ∆bfr mutant has a delay in morphological differentiation and pigmented antibiotic production (actinorhodin and undecylprodigiosin) on complex media. The effect of iron in minimal medium was also tested in the wild type and the ∆bfr mutant, and we observed different production of the two pigmented antibiotics in minimal medium, with differences between strains depending on iron concentration and the medium (solid or liquid). Contrary to expected, no intracellular iron differences were detected between the two strains, but there is a higher level of reactive oxygen species in the ∆bfr mutant and a higher tolerance to oxidative stress. Proteomics analysis showed no variation in iron response proteins and a lower abundance of proteins related to actinorhodin, ribosomal proteins and others related to secondary metabolism production and differentiation. On the contrary, it revealed a higher abundance of proteins related to different kind of stresses such as respiration and hypoxia, among others. The bacterioferritin of S. coelicolor is a new element in the complex regulation of the secondary metabolism in S. coelicolor and iron acts as a signal to modulate the biosynthesis of active molecules.
Project description:In order to define the impact of phosphate (Pi) availability on cellular metabolism the project aimed to perform a comparative analysis of the proteomes of two Streptomyces strains with different abilities to produce antibiotics, S. coelicolor and S. lividans as well as of the pptA mutant of S. lividans, grown low (1mM) and high (5mM) phosphate (Pi) availability conditions. Interestingly, in contrast to most Streptomyces species, S. coelicolor produces more antibiotics in Pi proficiency than in Pi limitation, S. lividans does not produce antibiotics in any Pi conditions and the pptA mutant produces antibiotics only in Pi limitation. This in-depth proteomic comparison of three Streptomyces strains (S. coelicolor, S. lividans wt and pptA mutant), in different growth conditions (time and Pi concentration in the medium) was performed on four biological replicates. Protein abundance changes were determined using two label-free mass spectrometry based-quantification methods: spectral count (SC) and MS1 ion intensities named XIC (for eXtracted Ion Current). Our proteomic data reveal for the first time, the impact of Pi availability on the abundance of approximately 4000 proteins of these Streptomyces strains with different abilities to produce antibiotics. The most striking feature differentiating these strains was the much higher abundance of enzymes of the respiratory chain in both phosphate conditions in S. coelicolor compared to the S. lividans strains.
Project description:This experiment aims to profile polyclonal antibody binding profiles in serum from vaccinated animals relative to antibody function in a virus neutralization assay. Rabbits received three vaccinations with a DNA vaccine encoding the spike protein of the SARS-CoV-2 index strain. Serum samples were selected based on a three-tier (low, intermediate, and high) capacity to cross-neutralize SARS-CoV-2 strains with known neutralization resistance. Following normalization of total anti-spike IgG levels, serum of each animal (n=3) were evaluated for antibody binding to 10mer cyclic constrained peptides spanning the entire spike protein and regions with known SARS-CoV-2 variant of concern spike mutations.
Project description:The impact on mRNA expression of the transcription factors Bas1, Pho2, Gcn4 and Gcr2p was investigated in the corresponding deletion strains during exponential growth in glucose minimal media batch cultures. All strains were grown in the same media and harvested during exponential growth, at the same cell density, for gene expression analysis. Gene expression analysis of each deletion mutant can be compared relative to the reference strain FY4. Three replicates each strain.
Project description:The impact on mRNA expression of the transcription factors Bas1, Pho2, Gcn4 and Gcr2p was investigated in the corresponding deletion strains during exponential growth in glucose minimal media batch cultures. All strains were grown in the same media and harvested during exponential growth, at the same cell density, for gene expression analysis. Gene expression analysis of each deletion mutant can be compared relative to the reference strain FY4. Three replicates each strain.
Project description:This study develops a pipeline for high-level production of the reverse antibiotic nybomycin from three seaweed species: Himanthalia elongata, Palmaria palmata, and Ulva lactuca. Screening Streptomyces strains identified S. explomaris, a marine species, as the best host to express the nybomycin gene cluster. The accumulated low yields in artificial seawater with brown seaweed hydrolysate. Gene expression analysis revealed downregulation of precursor supply pathways and upregulation of repressors, limiting biosynthesis. Metabolic engineering addressed these bottlenecks, leading to a superior S. explomaris mutant achieving 57 mg/L, a five-fold increase as compared to reported yields. The strain effectively valorized commercial seaweed hydrolysates, highlighting marine feedstocks' potential for antibiotic production.
Project description:RNA-seq was performed on T. maritima wild type, three glucose evolved cultures, and three glycerol adapted cultures. Wild type and glucose evolved strains were grown on glucose minimal media and glycerol evolved cultures were grown on glycerol minimal media. All samples were harvested in exponential phase.
Project description:The impact on mRNA expression of the transcription factors Bas1, Pho2, Gcn4 and Gcr2p was investigated in the corresponding deletion strains during exponential growth in glucose minimal media batch cultures. All strains were grown in the same media and harvested during exponential growth, at the same cell density, for gene expression analysis. Gene expression analysis of each deletion mutant can be compared relative to the reference strain FY4. Three replicates for wild-type and two replicates for the Pho2 deletion mutant.
Project description:The genomic and proteomic analyses of Streptomyces lividans strains deficient in the major signal peptidase SipY or in the translocase complex protein SecG resulted in a set of genes being equally regulated. These genes are apparently responsible for the common deficiencies in extracellular protein production and sporulation shared by both mutant strains, constituting a cellular response to the stress caused by the potential malfunction of the translocase complex, which we have named “extracellular protein translocation stress (EPTS)”.
Project description:The objective was to analyze the differential expression between the wild strain and the Streptomyces clavuligerus ΔclaR::aac mutant Six experimental conditions were assayed, two strains (Streptomyces clavuligerus ATCC 27064, S. clavuligerus ΔclaR::aac) in three culture times (22.5h, 46.5h and 60 h). Two biological replicates for each condition.