Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida
Sarcosine, an N-methylated amino acid, shows potential as antipsychotic, and serves as building block for peptide-based drugs, and acts as detergent when acetylated. N-methylated amino acids are mainly produced chemically or by biocatalysis, with either low yields or high costs for co-factor regener...
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Frontiers Media S.A.
2019-09-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/article/10.3389/fbioe.2019.00232/full |
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| author | Melanie Mindt Silvin Hannibal Maria Heuser Joe Max Risse Keerthi Sasikumar K. Madhavan Nampoothiri Volker F. Wendisch |
| author_facet | Melanie Mindt Silvin Hannibal Maria Heuser Joe Max Risse Keerthi Sasikumar K. Madhavan Nampoothiri Volker F. Wendisch |
| author_sort | Melanie Mindt |
| collection | DOAJ |
| description | Sarcosine, an N-methylated amino acid, shows potential as antipsychotic, and serves as building block for peptide-based drugs, and acts as detergent when acetylated. N-methylated amino acids are mainly produced chemically or by biocatalysis, with either low yields or high costs for co-factor regeneration. Corynebacterium glutamicum, which is used for the industrial production of amino acids for decades, has recently been engineered for production of N-methyl-L-alanine and sarcosine. Heterologous expression of dpkA in a C. glutamicum strain engineered for glyoxylate overproduction enabled fermentative production of sarcosine from sugars and monomethylamine. Here, mutation of an amino acyl residue in the substrate binding site of DpkA (DpkAF117L) led to an increased specific activity for reductive alkylamination of glyoxylate using monomethylamine and monoethylamine as substrates. Introduction of DpkAF117L into the production strain accelerated the production of sarcosine and a volumetric productivity of 0.16 g L−1 h−1 could be attained. Using monoethylamine as substrate, we demonstrated N-ethylglycine production with a volumetric productivity of 0.11 g L−1 h−1, which to the best of our knowledge is the first report of its fermentative production. Subsequently, the feasibility of using rice straw hydrolysate as alternative carbon source was tested and production of N-ethylglycine to a titer of 1.6 g L−1 after 60 h of fed-batch bioreactor cultivation could be attained. |
| first_indexed | 2024-12-13T05:54:33Z |
| format | Article |
| id | doaj.art-b1d9513da61349278e291cb8cd4271c4 |
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| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-12-13T05:54:33Z |
| publishDate | 2019-09-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-b1d9513da61349278e291cb8cd4271c42022-12-21T23:57:28ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852019-09-01710.3389/fbioe.2019.00232479143Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putidaMelanie Mindt0Silvin Hannibal1Maria Heuser2Joe Max Risse3Keerthi Sasikumar4K. Madhavan Nampoothiri5Volker F. Wendisch6Genetics of Prokaryotes, Faculty of Biology and CeBiTec, Bielefeld University, Bielefeld, GermanyGenetics of Prokaryotes, Faculty of Biology and CeBiTec, Bielefeld University, Bielefeld, GermanyGenetics of Prokaryotes, Faculty of Biology and CeBiTec, Bielefeld University, Bielefeld, GermanyFermentation Technology, Technical Faculty and CeBiTec, Bielefeld University, Bielefeld, GermanyMicrobial Processes and Technology Division, National Institute for Interdisciplinary Science and Technology, Council of Scientific & Industrial Research, Trivandrum, IndiaMicrobial Processes and Technology Division, National Institute for Interdisciplinary Science and Technology, Council of Scientific & Industrial Research, Trivandrum, IndiaGenetics of Prokaryotes, Faculty of Biology and CeBiTec, Bielefeld University, Bielefeld, GermanySarcosine, an N-methylated amino acid, shows potential as antipsychotic, and serves as building block for peptide-based drugs, and acts as detergent when acetylated. N-methylated amino acids are mainly produced chemically or by biocatalysis, with either low yields or high costs for co-factor regeneration. Corynebacterium glutamicum, which is used for the industrial production of amino acids for decades, has recently been engineered for production of N-methyl-L-alanine and sarcosine. Heterologous expression of dpkA in a C. glutamicum strain engineered for glyoxylate overproduction enabled fermentative production of sarcosine from sugars and monomethylamine. Here, mutation of an amino acyl residue in the substrate binding site of DpkA (DpkAF117L) led to an increased specific activity for reductive alkylamination of glyoxylate using monomethylamine and monoethylamine as substrates. Introduction of DpkAF117L into the production strain accelerated the production of sarcosine and a volumetric productivity of 0.16 g L−1 h−1 could be attained. Using monoethylamine as substrate, we demonstrated N-ethylglycine production with a volumetric productivity of 0.11 g L−1 h−1, which to the best of our knowledge is the first report of its fermentative production. Subsequently, the feasibility of using rice straw hydrolysate as alternative carbon source was tested and production of N-ethylglycine to a titer of 1.6 g L−1 after 60 h of fed-batch bioreactor cultivation could be attained.https://www.frontiersin.org/article/10.3389/fbioe.2019.00232/fullCorynebacterium glutamicumimine reductasemetabolic engineeringenzyme engineeringN-alkylated amino acidsN-methylamino acids |
| spellingShingle | Melanie Mindt Silvin Hannibal Maria Heuser Joe Max Risse Keerthi Sasikumar K. Madhavan Nampoothiri Volker F. Wendisch Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida Frontiers in Bioengineering and Biotechnology Corynebacterium glutamicum imine reductase metabolic engineering enzyme engineering N-alkylated amino acids N-methylamino acids |
| title | Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida |
| title_full | Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida |
| title_fullStr | Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida |
| title_full_unstemmed | Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida |
| title_short | Fermentative Production of N-Alkylated Glycine Derivatives by Recombinant Corynebacterium glutamicum Using a Mutant of Imine Reductase DpkA From Pseudomonas putida |
| title_sort | fermentative production of n alkylated glycine derivatives by recombinant corynebacterium glutamicum using a mutant of imine reductase dpka from pseudomonas putida |
| topic | Corynebacterium glutamicum imine reductase metabolic engineering enzyme engineering N-alkylated amino acids N-methylamino acids |
| url | https://www.frontiersin.org/article/10.3389/fbioe.2019.00232/full |
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