Streptomyces transglutaminase (TGase) is widely used to improve food texture properties. In this study, random mutagenesis and site-directed genetic modification were used to improve the production of TGase in Streptomyces mobaraensis. First, S. mobaraensis DSM40587 (smWT) was subjected to atmospheric and room-temperature plasma mutagenesis, and then a mutant (smY2019) with a 5.5-fold increase in TGase yield was screened from approximately 3000 × 25 (round) mutants. Compared to smWT, smY2019 exhibits a 3.2-fold higher TGase mRNA level and two site mutations within the -10 region of the TGase promoter. The recombinant expression analysis in the TGase-deficient S. mobaraensis suggests that the mutated TGase promoter is more robust than the wild-type one. Finally, we integrated two additional TGase expression cassettes into the smY2019 genome, yielding the recombinant strain smY2019-3C with a 103% increase in TGase production compared to smY2019. The smY2019-3C strain with 40 U/mL of TGase yield could be a suitable candidate for the industrial production of TGase.
Keywords: ARTP; Streptomyces mobaraensis; expression cassette; gene copy; promoter; transglutaminase.