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Antibiotic-free production of sucrose isomerase in Bacillus subtilis by genome integration.
- Source :
-
Biotechnology letters [Biotechnol Lett] 2024 Oct; Vol. 46 (5), pp. 781-789. Date of Electronic Publication: 2024 Jun 07. - Publication Year :
- 2024
-
Abstract
- Sucrose isomerase (SIase) catalyzes the hydrolysis and isomerization of sucrose to form isomaltulose, a valuable functional sugar widely used in the food industry. However, the lack of safe and efficient heterologous expression systems hinders SIase production and application. In this study, we achieved antibiotic-free SIase expression in Bacillus subtilis through genome integration. Using CRISPR/Cas9 system, SIase expression cassettes were integrated into various genomic loci, including amyE and ctc, both individually and in combination, resulting in single-copy and muti-copy integration strains. Engineered strains with a maltose-inducible promoter effectively expressed and secreted SIase. Notably, multi-copy strain exhibited enhanced SIase production, achieving 4.4 U/mL extracellular activity in shake flask cultivations. Furthermore, crude enzyme solution from engineered strain transformed high concentrations sucrose into high yields of isomaltulose, reaching a maximum yield of 94.6%. These findings demonstrate antibiotic-free SIase production in B. subtilis via genome integration, laying the foundation for its industrial production and application.<br /> (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)
- Subjects :
- Sucrose metabolism
Metabolic Engineering methods
CRISPR-Cas Systems genetics
Genome, Bacterial genetics
Bacterial Proteins genetics
Bacterial Proteins metabolism
Bacillus subtilis genetics
Bacillus subtilis enzymology
Isomaltose metabolism
Isomaltose analogs & derivatives
Glucosyltransferases genetics
Glucosyltransferases metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1573-6776
- Volume :
- 46
- Issue :
- 5
- Database :
- MEDLINE
- Journal :
- Biotechnology letters
- Publication Type :
- Academic Journal
- Accession number :
- 38847981
- Full Text :
- https://doi.org/10.1007/s10529-024-03501-3