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Improved Production of Pyrroloquinoline Quinone by Simultaneous Augmentation of Its Synthesis Gene Expression and Glucose Metabolism in Klebsiella pneumoniae

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Abstract

Klebsiella pneumoniae can naturally synthesize pyrroloquinoline quinone (PQQ), but current low yield restricts its commercialization. Here, we reported that PQQ production can be improved by simultaneously intensifying PQQ gene expression and glucose metabolism. Firstly, tandem repetitive tac promoters were constructed to overexpress PQQ synthesis genes. Results showed that when three repeats of tac promoter were recruited to overexpress PQQ synthesis genes, the recombinant strain generated 1.5-fold PQQ relative to the strain recruiting only one tac promoter. Quantitative real-time PCR (qRT-PCR) revealed the increased transcription levels of PQQ synthesis genes. Next, fermentation parameters were optimized to augment the glucose direct oxidation pathway (GDOP) mediated by PQQ-dependent glucose dehydrogenase (PQQ-GDH). Results demonstrated that the cultivation conditions of sufficient glucose (≥ 32 g/L), low pH (5.8), and limited potassium (0.7 nmol/L) significantly promoted the biosynthesis of gluconic acid, 2-ketogluconic acid, and PQQ. In optimum shake flask fermentation conditions, the K. pneumoniae strain overexpressing PQQ synthesis genes under three repeats of tac promoter generated 363.3 nmol/L of PQQ, which was 2.6-fold of that in original culture conditions. In bioreactor cultivation, this strain produced 2371.7 nmol/L of PQQ. To our knowledge, this is the highest PQQ titer reported so far using K. pneumoniae as a host strain. Overall, simultaneous intensification of pqq gene expression and glucose metabolism is effective to improve PQQ production.

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Acknowledgements

This work was supported by National Key Research and Development Program of China (2018YFA0901800), National Natural Science Foundation of China (21476011), and National High Technology Research and Development Program (863 Program) (2015AA021003).

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  1. College of Life Science and Technology, Beijing University of Chemical Technology, Beijing, 100029, People’s Republic of China

    Zhiwei Mi, Jingchao Cheng, Peng Zhao, Pingfang Tian & Tianwei Tan

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Mi, Z., Cheng, J., Zhao, P. et al. Improved Production of Pyrroloquinoline Quinone by Simultaneous Augmentation of Its Synthesis Gene Expression and Glucose Metabolism in Klebsiella pneumoniae. Curr Microbiol 77, 1174–1183 (2020). https://doi.org/10.1007/s00284-020-01918-3

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