Many Actinobacteria, most notably Streptomyces, produce structurally diverse bioactive natural products, including ribosomally synthesized peptides, by multistep enzymatic pathways. The use of site-specific genetic incorporation of unnatural amino acids to investigate and manipulate the functions of natural product biosynthetic enzymes, enzyme complexes, and ribosomally-derived peptides in these organisms would have important implications for drug discovery and development efforts. Here, we have designed, constructed, and optimized unnatural amino acid systems capable of incorporating p-iodo-l-phenylalanine and p-azido-l-phenylalanine site-specifically into proteins in the model natural product producer Streptomyces venezuelae ATCC 15439. We observed notable differences in the fidelity and efficiency of these systems between S. venezuelae and previously used hosts. Our findings serve as a foundation for using an expanded genetic code in Streptomyces to address questions related to natural product biosynthesis and mechanism of action that are relevant to drug discovery and development.
Summary
In this study, the evolutions of impedance and bio‐indicators (total aerobic count (TAC), total volatile base nitrogen (TVB‐N) and pH) of meat freshness during storage at 4 °C were investigated and compared. The influence of electrode configurations, frequencies and directions of applied electric fields on impedance measurement was considered. Good relationships between measured impedance and bio‐indicators were obtained, with R2 ranging from 0.758 to 0.992, RMSE from 0.11 to 0.57 for TAC and R2 ranging from 0.636 to 0.989, RMSE from 0.29 to 1.68 for TVB‐N. Bar and ring electrodes had better performance as compared to needle electrodes. Effects of frequency on impedance were important from 20 Hz to 200 kHz and diminished in the higher ranges. Impedance generally exhibited better correlations with TAC, TVB‐N at high frequency than at low frequency. The findings demonstrated that impedance measurement could be a viable, rapid method to assess the quality of stored pork meat.
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