Evaluation of an oxygen‐dependent self‐inducible surfactin synthesis in B. subtilis by substitution of native promoter P srfA by anaerobically active P narG and P nasD
Mareen Hoffmann,
Alina Braig,
Diana Stephanie Fernandez Cano Luna,
Katharina Rief,
Philipp Becker,
Chantal Treinen,
Peter Klausmann,
Kambiz Morabbi Heravi,
Marius Henkel,
Lars Lilge,
Rudolf Hausmann
Affiliations
Mareen Hoffmann
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Alina Braig
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Diana Stephanie Fernandez Cano Luna
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Katharina Rief
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Philipp Becker
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Chantal Treinen
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Peter Klausmann
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Kambiz Morabbi Heravi
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Marius Henkel
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Lars Lilge
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Rudolf Hausmann
Institute of Food Science and Biotechnology (150), Department of Bioprocess Engineering (150k), University of Hohenheim
Abstract A novel approach targeting self-inducible surfactin synthesis under oxygen-limited conditions is presented. Because both the nitrate (NarGHI) and nitrite (NasDE) reductase are highly expressed during anaerobic growth of B. subtilis, the native promoter P srfA of the surfactin operon in strain B. subtilis JABs24 was replaced by promoters P narG and P nasD to induce surfactin synthesis anaerobically. Shake flask cultivations with varying oxygen availabilities indicated no significant differences in native P srfA expression. As hypothesized, activity of P narG and P nasD increased with lower oxygen levels and surfactin was not produced by P srfA ::P narG as well as P srfA ::P nasD mutant strains under conditions with highest oxygen availability. P narG showed expressions similar to P srfA at lowest oxygen availability, while maximum value of P nasD was more than 5.5-fold higher. Although the promoter exchange P srfA ::P narG resulted in a decreased surfactin titer at lowest oxygen availability, the strain carrying P srfA ::P nasD reached a 1.4-fold increased surfactin concentration with 696 mg/L and revealed an exceptional high overall Y P/X of 1.007 g/g. This value also surpassed the Y P/X of the reference strain JABs24 at highest and moderate oxygen availability. Bioreactor cultivations illustrated that significant cell lysis occurred when the process of “anaerobization” was performed too fast. However, processes with a constantly low agitation and aeration rate showed promising potential for process improvement, especially by employing the strain carrying P srfA ::P nasD promoter exchange. Additionally, replacement of other native promoters by nitrite reductase promoter P nasD represents a promising tool for anaerobic-inducible bioprocesses in Bacillus.
