Streptomyces rimosus is an industrially important microorganism with efficient production of secondary metabolites. This strain gained increased interest as a host for heterologous protein biosynthesis. The aim of this master’s thesis was the development and optimization of an oksitetracicline (OTC)-inducible promoter system Potr in S. rimosus strains lacking the OTC biosynthetic pathway. We applyied reporter gene mCherry to monitor protein expression by measuring fluorescence. In the course of experiments, different concentrations of OTC inducer at different induction time points were tested to evaluate their impact on culture growth and the amount of heterologous protein produced. We observed that the optimal inducer concentration ranged between 50 and 250 µM, with the highest expression achieved when induction was carried between the 2. and 12. hour after the inoculation of production phase. Strains with an additional 145 kb genomic deletion exhibited increased heterologous protein production, suggesting that the reduction of metabolic resources was favourable for heterologous expression. The highest concentration of the heterologous protein reached up to 22 mg/L (± 2 mg/L). Strains without OTC biosynthetic gene cluster as well as additional 145 kb genomic deletion exhibited the highest capacity for protein secretion. Analysis of protein secretion revealed that approximately 80% of the heterologous protein was secreted, indicating efficient functioning of the Sec secretion pathway.
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