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Efficient biosynthesis of D/L-alanine in the recombinant Escherichia coli BL21(DE3) by biobrick approach
by
Chu, Mengqiu
, Huang, Xinyan
, Hao, Shimiao
, Zhang, Xuan
, He, Guangzheng
, Zhao, Baohua
, Naeem, Muhammad
, Ju, Jiansong
, Wang, Jiaying
in
Acidification
/ Alanine dehydrogenase
/ Alanine racemase
/ Amino acids
/ Ammonium chloride
/ Antibiotics
/ Bacteria
/ biobrick method
/ Bioengineering and Biotechnology
/ Biosynthesis
/ Cloning
/ D-Alanine
/ D-glucose
/ D/L-alanine
/ Dehydrogenases
/ E coli
/ Enzymes
/ Escherichia coli
/ Fermentation
/ Food industry
/ Gene expression
/ Genetic engineering
/ Glucose
/ Glucose dehydrogenase
/ L-Alanine
/ Methods
/ Microorganisms
/ optimization
/ Oxidation
/ Plasmids
/ Polyesters
/ Protein expression
/ Proteins
/ Pyruvic acid
/ SDS-PAGE
/ Sodium bicarbonate
/ Sodium carbonate
/ Sodium pyruvate
/ Tuberculosis
2024
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Efficient biosynthesis of D/L-alanine in the recombinant Escherichia coli BL21(DE3) by biobrick approach
by
Chu, Mengqiu
, Huang, Xinyan
, Hao, Shimiao
, Zhang, Xuan
, He, Guangzheng
, Zhao, Baohua
, Naeem, Muhammad
, Ju, Jiansong
, Wang, Jiaying
in
Acidification
/ Alanine dehydrogenase
/ Alanine racemase
/ Amino acids
/ Ammonium chloride
/ Antibiotics
/ Bacteria
/ biobrick method
/ Bioengineering and Biotechnology
/ Biosynthesis
/ Cloning
/ D-Alanine
/ D-glucose
/ D/L-alanine
/ Dehydrogenases
/ E coli
/ Enzymes
/ Escherichia coli
/ Fermentation
/ Food industry
/ Gene expression
/ Genetic engineering
/ Glucose
/ Glucose dehydrogenase
/ L-Alanine
/ Methods
/ Microorganisms
/ optimization
/ Oxidation
/ Plasmids
/ Polyesters
/ Protein expression
/ Proteins
/ Pyruvic acid
/ SDS-PAGE
/ Sodium bicarbonate
/ Sodium carbonate
/ Sodium pyruvate
/ Tuberculosis
2024
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Efficient biosynthesis of D/L-alanine in the recombinant Escherichia coli BL21(DE3) by biobrick approach
by
Chu, Mengqiu
, Huang, Xinyan
, Hao, Shimiao
, Zhang, Xuan
, He, Guangzheng
, Zhao, Baohua
, Naeem, Muhammad
, Ju, Jiansong
, Wang, Jiaying
in
Acidification
/ Alanine dehydrogenase
/ Alanine racemase
/ Amino acids
/ Ammonium chloride
/ Antibiotics
/ Bacteria
/ biobrick method
/ Bioengineering and Biotechnology
/ Biosynthesis
/ Cloning
/ D-Alanine
/ D-glucose
/ D/L-alanine
/ Dehydrogenases
/ E coli
/ Enzymes
/ Escherichia coli
/ Fermentation
/ Food industry
/ Gene expression
/ Genetic engineering
/ Glucose
/ Glucose dehydrogenase
/ L-Alanine
/ Methods
/ Microorganisms
/ optimization
/ Oxidation
/ Plasmids
/ Polyesters
/ Protein expression
/ Proteins
/ Pyruvic acid
/ SDS-PAGE
/ Sodium bicarbonate
/ Sodium carbonate
/ Sodium pyruvate
/ Tuberculosis
2024
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Efficient biosynthesis of D/L-alanine in the recombinant Escherichia coli BL21(DE3) by biobrick approach
Journal Article
Efficient biosynthesis of D/L-alanine in the recombinant Escherichia coli BL21(DE3) by biobrick approach
2024
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Overview
Alanine is the most abundant chiral amino acid that exists into the D -alanine or L-alanine forms with diverse applications in the biomedical, pharmaceutical, plastics, and food industries. D/L-alanine production can be carried out through chemical, microbial fermentation, and biocatalytic methods and not much effective due to complicated processes or purification issues and is still challenging to achieve a higher yield. In the present study, biobrick method was utilized for efficient production of D/L-alanine in the recombinant Escherichia coli BL21(DE3) with tandem three-gene co-expression plasmid. Firstly, the co-expression plasmid pET-22bNS-DadX-Ald-Gdh containing three genes, alanine dehydrogenase ( ald) , alanine racemase ( dadX) , and glucose dehydrogenase ( gdh) from Bacillus pseudofirmus OF4 were successfully constructed and introduced into the E. coli BL21(DE3) strain. Then, under optimized conditions in the whole-cell biocatalytic reaction [20 mM Na 2 CO 3 -NaHCO 3 (pH 10.1), 200 mM D-glucose, 200 mM sodium pyruvate, and 200 mM ammonium chloride], the concentration of D-alanine and L-alanine reached the maximum value (6.48 g/L and 7.05 g/L) after 3.0 h reaction time at 37°C under 180 rpm rotation. Meanwhile, promoter replacement experiments and Western blot analysis revealed that the expression level of protein OF4Ald had a significant effect on the production of D/L-alanine, indicating that alanine dehydrogenase might be the rate-limiting enzyme for D/L-alanine synthesis. This study provides a simple, feasible, and efficient biosynthesis process of D/L-alanine, which could explore emerging applications for large-scale production of industrial bioproducts.
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