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Development of a potential stationary-phase specific gene expression system by engineering of SigB-dependent cg3141 promoter in Corynebacterium glutamicum

  • Applied genetics and molecular biotechnology
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Abstract

Corynebacterium glutamicum is a non-pathogenic, non-sporulating Gram-positive soil bacterium that has been used for the industrial production of various proteins and chemicals. To achieve enhanced and economical production of target molecules, the development of strong auto-inducible promoters is desired, which can be activated without expensive inducers and has significant advantages for industrial-scale use. Here, we developed a stationary-phase gene expression system by engineering a sigma factor B (SigB)-dependent promoter that can be activated during the transition phase between exponential and stationary growth phases in C. glutamicum. First, the inducibilities of three well-known SigB-dependent promoters were examined using super-folder green fluorescent protein as a reporter protein, and we found that promoter of cg3141 (P cg3141 ) exhibited the highest inducibility. Next, a synthetic promoter library was constructed by randomizing the flanking and space regions of P cg3141 , and the stationary-phase promoters exhibiting high strengths were isolated via FACS-based high-throughput screening. The isolated synthetic promoter (P4-N14) showed a 3.5-fold inducibility and up to 20-fold higher strength compared to those of the original cg3141 promoter. Finally, the use of the isolated P4-N14 for fed-batch cultivation was verified with the production of glutathione S-transferase as a model protein in a lab-scale (5-L) bioreactor.

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Acknowledgments

This work was supported by the Intelligent Synthetic Biology Center of Global Frontier Project (Grant no. NRF-2014M3A6A8066443) and by the National Research Foundation of Korea (Grant no. NRF-2015R1A2A2A01007674) funded by the Ministry of Science, ICT and Future Planning (MSIP).

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Authors and Affiliations

  1. Department of Chemical and Biomolecular Engineering, BK21 Plus Program, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 34141, Republic of Korea

    Min Jeong Kim, Sung Sun Yim, Jae Woong Choi & Ki Jun Jeong

  2. Institute for the BioCentury, KAIST, 291 Daehak-ro, Yuseong-gu, Daejeon, 34141, Republic of Korea

    Ki Jun Jeong

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  1. Min Jeong Kim
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  2. Sung Sun Yim
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Correspondence to Ki Jun Jeong.

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Kim, M.J., Yim, S.S., Choi, J.W. et al. Development of a potential stationary-phase specific gene expression system by engineering of SigB-dependent cg3141 promoter in Corynebacterium glutamicum . Appl Microbiol Biotechnol 100, 4473–4483 (2016). https://doi.org/10.1007/s00253-016-7297-y

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  • DOI: https://doi.org/10.1007/s00253-016-7297-y

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