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Article
2025
Recombinant production of acidophilic L-arabinose isomerase from Lentilactobacillus parakefiri in Bacillus subtilis
Recombinant production of acidophilic L-arabinose isomerase from Lentilactobacillus parakefiri in Bacillus subtilis
Abstract (English)
Background: The monosaccharide D-tagatose is a promising alternative to sucrose because of its similar sweetness and lower glycemic index. A novel L-arabinose isomerase (L-AI) from Lentilactobacillus parakefiri DSM 10551 (L-AI-Lp) has been biochemically characterized and used to isomerize D-galactose to D-tagatose in skim milk ultrafiltration permeate at pH 4.5 and 6.5. However, like most L-AIs described in the literature, this enzyme has only been produced recombinantly in Escherichia coli. This study aimed to systematically investigate the intracellular recombinant production of L-AI-Lp in Bacillus subtilis, which has qualified for a presumption of safety (QPS) designation from the European Food Safety Authority. Results: The influence of four promoters on L-AI-Lp production in B. subtilis 007 was investigated in shake flask cultivations. Among these, the PAprE promoter yielded the highest volumetric L-AI activity of 69.2 ± 7.4 µkatGal, 65 °C/LCulture. The production yield was further increased to 147.7 ± 1.0 µkatGal, 65 °C/LCulture by using the nonsporulating, surfactin-deficient strain B. subtilis 007 ∆sfp ∆sigF, which was constructed by deleting sigF and sfp in B. subtilis 007. Furthermore, the influence of pH and dissolved oxygen (DO) on bioreactor cultivations of B. subtilis 007 ∆sfp ∆sigF was analyzed. In bioreactor cultivations, the highest L-AI activity of 88.6 ± 2.4 µkatGal, 65 °C/LCulture was measured under unregulated pH and low oxygen conditions (DO ≤ 5%), representing a 3.2-fold increase compared with previous recombinant production in E. coli. The L-AI-Lp was subsequently partially purified by heat treatment and precipitation methods, resulting in a 7.8-fold increase in specific activity to 128.2 nkatGal, 65 °C/mg and a yield of 84%. Conclusions: The L-AI-Lp was recombinantly produced for the first time in a microbial species with QPS status using the nonsporulating and surfactin-deficient strain B. subtilis 007 ∆sfp ∆sigF. The L-AI-Lp was subsequently partially purified via nonchromatographic methods, providing a basis for a low-cost downstream process. These results represent an important step toward potential industrial application of L-AI-Lp and highlight the potential of B. subtilis 007 ∆sfp ∆sigF as an expression host for the recombinant production of L-AIs compared with previously used hosts from the order Lactobacillales.
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Microbial cell factories, 24 (2025), 1, 251.
https://doi.org/10.1186/s12934-025-02900-z.
ISSN: 1475-2859
London : BioMed Central
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Weber, N., Götz, S., Senger, J., Lutz-Wahl, S., & Fischer, L. (2025). Recombinant production of acidophilic L-arabinose isomerase from Lentilactobacillus parakefiri in Bacillus subtilis. Microbial cell factories, 24(1). https://doi.org/10.1186/s12934-025-02900-z
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English
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660 Chemical engineerin
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University bibliography
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Sustainable Development Goals
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@article{Weber2025,
doi = {10.1186/s12934-025-02900-z},
author = {Weber, Nathanael and Götz, Sebastian and Senger, Jana et al.},
title = {Recombinant production of acidophilic L-arabinose isomerase from Lentilactobacillus parakefiri in Bacillus subtilis},
journal = {Microbial Cell Factories},
year = {2025},
volume = {24},
number = {1},
}