Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production
Abstract Background Alanine dehydrogenase (AlaDH) belongs to oxidoreductases, and it exists in several different bacteria species and plays a key role in microbial carbon and nitrogen metabolism, spore formation and photosynthesis. In addition, AlaDH can also be applied in biosynthesis of L-alanine...
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| Format: | Article |
| Language: | English |
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BMC
2023-08-01
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| Series: | Biotechnology for Biofuels and Bioproducts |
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| Online Access: | https://doi.org/10.1186/s13068-023-02373-5 |
| _version_ | 1797453557743484928 |
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| author | Pengfei Gu Qianqian Ma Shuo Zhao Qiang Li Juan Gao |
| author_facet | Pengfei Gu Qianqian Ma Shuo Zhao Qiang Li Juan Gao |
| author_sort | Pengfei Gu |
| collection | DOAJ |
| description | Abstract Background Alanine dehydrogenase (AlaDH) belongs to oxidoreductases, and it exists in several different bacteria species and plays a key role in microbial carbon and nitrogen metabolism, spore formation and photosynthesis. In addition, AlaDH can also be applied in biosynthesis of L-alanine from cheap carbon source, such as glucose. Results To achieve a better performance of L-alanine accumulation, system evaluation and comparison of different AlaDH with potential application value are essential. In this study, enzymatic properties of AlaDH from Bacillus subtilis 168 (BsAlaDH), Bacillus cereus (BcAlaDH), Mycobacterium smegmatis MC2 155 (MsAlaDH) and Geobacillus stearothermophilus (GsAlaDH) were firstly carefully investigated. Four different AlaDHs have few similarities in optimum temperature and optimum pH, while they also exhibited significant differences in enzyme activity, substrate affinity and enzymatic reaction rate. The wild E. coli BL21 with these four AlaDHs could produce 7.19 g/L, 7.81 g/L, 6.39 g/L and 6.52 g/L of L-alanine from 20 g/L glucose, respectively. To further increase the L-alanine titer, competitive pathways for L-alanine synthesis were completely blocked in E. coli. The final strain M-6 could produce 80.46 g/L of L-alanine with a yield of 1.02 g/g glucose after 63 h fed-batch fermentation, representing the highest yield for microbial L-alanine production. Conclusions Enzyme assay, biochemical characterization and structure analysis of BsAlaDH, BcAlaDH, MsAlaDH and GsAlaDH were carried out. In addition, application potential of these four AlaDHs in L-alanine productions were explored. The strategies here can be applied for developing L-alanine producing strains with high titers. |
| first_indexed | 2024-03-09T15:24:32Z |
| format | Article |
| id | doaj.art-9dc0d70757cc46669fd309b984a4dd0e |
| institution | Directory Open Access Journal |
| issn | 2731-3654 |
| language | English |
| last_indexed | 2024-03-09T15:24:32Z |
| publishDate | 2023-08-01 |
| publisher | BMC |
| record_format | Article |
| series | Biotechnology for Biofuels and Bioproducts |
| spelling | doaj.art-9dc0d70757cc46669fd309b984a4dd0e2023-11-26T12:36:08ZengBMCBiotechnology for Biofuels and Bioproducts2731-36542023-08-0116111610.1186/s13068-023-02373-5Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine productionPengfei Gu0Qianqian Ma1Shuo Zhao2Qiang Li3Juan Gao4School of Biological Science and Technology, University of JinanSchool of Biological Science and Technology, University of JinanSchool of Biological Science and Technology, University of JinanSchool of Biological Science and Technology, University of JinanSchool of Biological Science and Technology, University of JinanAbstract Background Alanine dehydrogenase (AlaDH) belongs to oxidoreductases, and it exists in several different bacteria species and plays a key role in microbial carbon and nitrogen metabolism, spore formation and photosynthesis. In addition, AlaDH can also be applied in biosynthesis of L-alanine from cheap carbon source, such as glucose. Results To achieve a better performance of L-alanine accumulation, system evaluation and comparison of different AlaDH with potential application value are essential. In this study, enzymatic properties of AlaDH from Bacillus subtilis 168 (BsAlaDH), Bacillus cereus (BcAlaDH), Mycobacterium smegmatis MC2 155 (MsAlaDH) and Geobacillus stearothermophilus (GsAlaDH) were firstly carefully investigated. Four different AlaDHs have few similarities in optimum temperature and optimum pH, while they also exhibited significant differences in enzyme activity, substrate affinity and enzymatic reaction rate. The wild E. coli BL21 with these four AlaDHs could produce 7.19 g/L, 7.81 g/L, 6.39 g/L and 6.52 g/L of L-alanine from 20 g/L glucose, respectively. To further increase the L-alanine titer, competitive pathways for L-alanine synthesis were completely blocked in E. coli. The final strain M-6 could produce 80.46 g/L of L-alanine with a yield of 1.02 g/g glucose after 63 h fed-batch fermentation, representing the highest yield for microbial L-alanine production. Conclusions Enzyme assay, biochemical characterization and structure analysis of BsAlaDH, BcAlaDH, MsAlaDH and GsAlaDH were carried out. In addition, application potential of these four AlaDHs in L-alanine productions were explored. The strategies here can be applied for developing L-alanine producing strains with high titers.https://doi.org/10.1186/s13068-023-02373-5Alanine dehydrogenaseL-alanineEscherichia coliMetabolic engineering |
| spellingShingle | Pengfei Gu Qianqian Ma Shuo Zhao Qiang Li Juan Gao Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production Biotechnology for Biofuels and Bioproducts Alanine dehydrogenase L-alanine Escherichia coli Metabolic engineering |
| title | Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production |
| title_full | Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production |
| title_fullStr | Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production |
| title_full_unstemmed | Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production |
| title_short | Alanine dehydrogenases from four different microorganisms: characterization and their application in L-alanine production |
| title_sort | alanine dehydrogenases from four different microorganisms characterization and their application in l alanine production |
| topic | Alanine dehydrogenase L-alanine Escherichia coli Metabolic engineering |
| url | https://doi.org/10.1186/s13068-023-02373-5 |
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