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High yield secretion of heterologous proteins in Corynebacterium glutamicum using its own Tat-type signal sequence

  • Applied Genetics and Molecular Biotechnology
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Abstract

Efficient protein secretion, the basis of large-scale production of many compounds central to the biotechnology industry, is achieved by signal peptide and propeptide optimization in addition to optimizing host factors affecting heterologous protein production. Here, we fused green fluorescent protein (GFP) to the recently identified Tat-type secretory signal peptide of CgR0949 to demonstrate a high-yield protein secretion system of Corynebacterium glutamicum. The resultant secretion vector facilitated effective secretion of active-form GFP (20 mg l−1) into C. glutamicum culture medium. The expression of GFP was enhanced 2.9-fold using the Shine–Dalgarno sequence of triosephosphate isomerase in the secretion vector. Moreover, GFP drastically accumulated in the culture supernatant upon addition of calcium chloride even though Ca2+ addition did neither enhanced the transcription of gfp nor resulted in the accumulation of cytosolic GFP. Active-form GFP concentration reached 1.8 g l−1 after 48-h incubation in a jar fermentor. Likewise, α-amylase accumulation in C. glutamicum cultures was also enhanced by Ca2+ addition, suggesting that Ca2+ may affect general protein secretion in C. glutamicum.

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Acknowledgment

We thank Dr. C. Omumasaba (internal) for critical reading of the manuscript. This study was partly funded by the New Energy and industrial Technology Development Organization (NEDO).

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Authors and Affiliations

  1. Molecular Microbiology and Biotechnology Group, Research Institute of Innovative Technology for the Earth (RITE), 9-2 Kizugawadai, Kizugawa, Kyoto, 619-0292, Japan

    Hiroshi Teramoto, Keiro Watanabe, Nobuaki Suzuki, Masayuki Inui & Hideaki Yukawa

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  1. Hiroshi Teramoto
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  2. Keiro Watanabe
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  3. Nobuaki Suzuki
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  4. Masayuki Inui
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  5. Hideaki Yukawa
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Correspondence to Hideaki Yukawa.

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Teramoto, H., Watanabe, K., Suzuki, N. et al. High yield secretion of heterologous proteins in Corynebacterium glutamicum using its own Tat-type signal sequence. Appl Microbiol Biotechnol 91, 677–687 (2011). https://doi.org/10.1007/s00253-011-3281-8

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  • DOI: https://doi.org/10.1007/s00253-011-3281-8

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