Project description:Comparative genomic hybridization analysis of Streptomyces coelicolor A3(2) versus Streptomyces lividans 66 and Streptomyces lividans TK24 using high density 105,000 x 60-mer ink-jet in situ synthesized arrays.
Project description:By the direct comparison of illuminated and non-illuminated microtiter plate cultures of Streptomyces lividans TK24 transcriptomes, the effects of differing cultivation systems by the relevance of light, a parameter that is usually out of scope during heterotrophic bioprocesses, could be addressed.
Project description:The Streptomyces lividans lsp gene encodes a type II signal peptidase (Lsp) that cleaves the type II leader peptides of lipoproteins. Transcriptional profiling of the bacterium depleted of the lsp gene mainly resulted in deactivation of the sigma U regulon, as well as in downregulation of genes involved in the biogenesis and function of ribosomes and genes encoding some major secretory proteins as determined by hybridisation of commercially available S. lividans genome-wide microarrays. Almost 50% of the dowregulated genes have been described as forming part of the stringent response in streptomycetes. The gene encoding the S. lividans extracellular foldase, the lipoprotein FkpA, is equally downregulated. Therefore, the deletion of lsp from the S. livdans genome temporarily triggers a cellular stress where the stringent response is, at least, partially induced.
Project description:When SCO0988 was overexpressed in Streptomyces lividans TK24, it reveals the mechanism of acetyltransferase SCO0988 on regulating secondary metabolism by comparing acetylome of overexpressed strains and control strains
Project description:The gene aml encoding alpha-amylase in Streptomyces lividans was cloned in the multicopy plasmid pIJ486, generating plasmid pAMI11. Plasmid pAMI11 and pIJ486 were propagated in S. lividans TK21 to obtain S. lividans TK21(pAMI11) and its isogenic strain S. lividans TK21(pIJ486). Transcriptional profiling of the bacterium that overproduces alpha-amylase mainly resulted in the upregulation of genes involved in the biogenesis and function of ribosomes, together with the upregulation of the genes involved in the redox processes, the ABC transporters, the central carbon, aminoacid and purine /pyrimidine metabolism. Moreover, some genes involved in oxidative stress were upregulated. The number of genes downregulated was much lower than the upregulated ones. Therefore, bacteria respond by favouring alpha-amylase overproduction that apparently does not cause damage to the cell.
Project description:S. lividans TK24 is a popular host for the production of small molecules and for the secretion of heterologous proteins. TK24 has a large genome with at least 29 secondary metabolite gene clusters that are non-essential for viability and undergo complex regulation. To optimize heterologous protein secretion, we previously constructed ten chassis strains that are devoid of several secondary metabolite gene clusters. Genome reduction was aimed at reducing carbon flow to secondary metabolites and pigmentation in the spent growth medium and improving colony morphology. Strains RG1.0-RG1.10 contain various deletion combinations of the blue actinorhodin cluster (act), the calcium-dependent antibiotic (cda), the undecylprodigiosin (red) and coelimycin A (cpk) clusters, the melanin cluster (mel), the matAB genes that affect mycelial aggregation and the non-essential sigma factor hrdD that controls the transcription of Act and Red regulatory proteins. Two derivative strains, RG1.5 and 1.9, showed a ~15% reduction in growth rate, >2-fold increase in the total mass yield of their native secretome and altered abundance of several specific proteins compared with TK24. Metabolomics and RNAseq analysis revealed that genome reduction led to rapid cessation of growth due to aminoacid depletion and caused both redox and cell envelope stresses, upregulation of the Sec-pathway components secDF and chaperones and a cell envelope two component regulator. RG1.9 maintained elevated heterologous secretion of mRFP and mTNFα by 12-70%. An integrated model is presented linking genome reduction and enhanced secretion. Importance: S. lividans TK24 encode 29 secondary metabolite gene clusters controlled with highly complex systems. This study established an important step toward understanding how secondary metabolite clusters can be manipulated to construct a surrogate TK24 platform with optimized metabolite funnelling. Using integrative multi-omics tools with protein secretion we provide an in-depth view of this fascinating complex process and its mechanistic regulation.