Cloned (Comment) | Organism |
---|---|
gene thrA, recombinant overexpression of enzyme mutant G433R in Escherichia coli strain Bl21 | Escherichia coli |
Protein Variants | Comment | Organism |
---|---|---|
G433R | site-directed mutagenesis, strain HS33/pACYC-pycP458S-thrAG433R-lysC shows increased activity (62.4% of the maximum theoretical yield) | Escherichia coli |
additional information | key metabolic pathway for construction of an inducer-free L-homoserine-producing strain to maximize the productivity of L-homoserine based on genetic-engineering tools, comparison of L-homoserine production, cell growth, and glucose consumption in different engineered strains, overview. L-Homoserine is a nonessential amino acid for the biosynthesis of L-threonine and L-methionine. It is also an important precursor for the production of isobutanol, 1,4-butanediol, L-phosphinothricin, 2,4-ihydroxybutyrate, and 1,3-propanediol. The initial L-homoserine-producing strain HS1 is obtained by blocking the degradative and competitive pathways and overexpressing thrA (encoding homoserine dehydrogenase) based on an O-succinyl homoserine-producing strain, using the pull-push-block strategy, an efficient method to engineer microorganisms involved in biosynthesizing target products by modifying metabolic networks. L-homoserine-converting pathway-related genes (thrB, encoding homoserine kinase, and metA, encoding homoserine O-succinyltransferase) are successively deleted to block L-homoserine degradation. Gene thrA is overexpressed to push the carbon flux to L-homoserine production. Then, the lysine-auxotrophic strain HS2 is generated by deleting lysA to eliminate a precursor competing metabolic pathway on L-homoserine production. For strengthening the capability of the L-homoserine transport system and the transformation of other toxic intermediate metabolites, gene rhtA, encoding the inner membrane transporter that is involved in the export of L-homoserine, is overexpressed chromosomally by replacing the native promoter with the trc promoter to obtain strain HS3 (Trc-rhtA). The strain shows increased activity. Increase in the L-homoserine export capacity and relieve the growth burden of homoserine-producing strains to enable survival via replacement of the native promoter of the eamA gene by the trc promoter in strain HS4 (Trc-eamA). Two rhtA gene copies (the native rhtA gene and replacement of the lacI gene) and eamA are overexpressed under the trc promoter in the chromosome to construct strain HS5 (DELTAlacI::Trc-rhtA Trc-rhtA Trc-eamA). Under batch culture, strain HS5, with modification of the transport system and construction of a constitutive expression system, can produce 3.14 g/l L-homoserine, which is 54.2% higher than strain HS2 production. In addition, the specific production of strain HS5 is also increased. Repression of candidate genes by the CRISPRi system to further enhance L-homoserine production | Escherichia coli |
P458S | site-directed mutagenesis, strain HS33/pACYC-pycP458S-thrAG433R-lysC shows increased activity (62.4% of the maximum theoretical yield) | Escherichia coli |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
additional information | L-homoserine inhibits the activity of aspartokinase encoded by metL | Escherichia coli |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-homoserine + NAD+ | Escherichia coli | - |
L-aspartate 4-semialdehyde + NADH + H+ | - |
? | |
L-homoserine + NAD+ | Escherichia coli W3110 | - |
L-aspartate 4-semialdehyde + NADH + H+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Escherichia coli | P00561 | - |
- |
Escherichia coli W3110 | P00561 | - |
- |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
L-homoserine + NAD+ | - |
Escherichia coli | L-aspartate 4-semialdehyde + NADH + H+ | - |
? | |
L-homoserine + NAD+ | - |
Escherichia coli W3110 | L-aspartate 4-semialdehyde + NADH + H+ | - |
? | |
additional information | the bifunctional enzyme also exhibits aspartokinase activity, EC 2.7.2.4 | Escherichia coli | ? | - |
- |
|
additional information | the bifunctional enzyme also exhibits aspartokinase activity, EC 2.7.2.4 | Escherichia coli W3110 | ? | - |
- |
Synonyms | Comment | Organism |
---|---|---|
More | see also EC 2.7.2.4 | Escherichia coli |
thrA | - |
Escherichia coli |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
NAD+ | - |
Escherichia coli | |
NADH | - |
Escherichia coli |