Abstract
Objectives
To improve target protein production by manipulating expression levels of alanine racemase in Bacillus licheniformis.
Results
The gene of dal was identified to be responsible for alanine racemase function. Based on the selection marker of dal, a food-grade expression system was constructed in B. licheniformis, and effects of different dal expression levels mediated by promoters on α-amylase production were investigated. The highest α-amylase activity (155 U/ml) was obtained in BL10D/pP43SAT-PtetDal, increased by 27% compared with that of the control strain BL10/pP43SAT in tetracycline-based system (123 U/ml). Moreover, the dal transcriptional level was not correlated positively with that of amyL.
Conclusions
A food-grade system for high-level production of α-amylase was constructed in B. licheniformis, revealing that expression levels of selection marker significantly affected target protein production.
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Acknowledgements
This work was supported by the National Science & Technology Pillar Program during the twelfth five-year plan period (2013AA102801-52), the Science and Technology Program of Wuhan (20160201010086).
Supporting information
Supplementary Table 1—The strains and plasmids used in this study.
Supplementary Table 2—The primers used in this research.
Supplementary Fig. 1—Confirmation of gene deficient strains by PCR amplification.
Supplementary Fig. 2—Effects of dal or alrB deficiency on the cell growth in the medium with or without d-alanine.
Supplementary Fig. 3—The flow chart for construction of the co-expression vectors.
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He, P., Zhang, Z., Cai, D. et al. High-level production of α-amylase by manipulating the expression of alanine racamase in Bacillus licheniformis . Biotechnol Lett 39, 1389–1394 (2017). https://doi.org/10.1007/s10529-017-2359-5
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DOI: https://doi.org/10.1007/s10529-017-2359-5