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Literature summary extracted from

  • Weiss, M.S.; Pavlidis, I.V.; Spurr, P.; Hanlon, S.P.; Wirz, B.; Iding, H.; Bornscheuer, U.T.
    Amine transaminase engineering for spatially bulky substrate acceptance (2017), ChemBioChem, 18, 1022-1026 .
    View publication on PubMed

Application

EC Number Application Comment Organism
2.6.1.B16 synthesis amine transaminase (ATA) catalyzing stereoselective amination of prochiral ketones is an attractive alternative to transition metal catalysis Ruegeria sp. TM1040
2.6.1.B21 synthesis amine transaminase (ATA) catalyzing stereoselective amination of prochiral ketones is an attractive alternative to transition metal catalysis Ruegeria sp. TM1040

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.6.1.B21 gene pabC, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, recombinant expression of His-tagged enzyme in Escherichia coli strain BL21(DE3) Ruegeria sp. TM1040

Protein Variants

EC Number Protein Variants Comment Organism
2.6.1.B16 additional information mutational analysis of enzyme-substrate interactions and substrate specificity, overview Ruegeria sp. TM1040
2.6.1.B16 P281S site-directed mutagenesis, the mutation seems to have a negative effect on specific activity Ruegeria sp. TM1040
2.6.1.B16 Y152F site-directed mutagenesis, the mutation stabilizes the enzyme, the activity is slightly reduced compared to wild-type Ruegeria sp. TM1040
2.6.1.B16 Y59W/T231A site-directed mutagenesis, inactive mutant Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87F/T231A site-directed mutagenesis, inactive mutant Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87L/T231A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87L/T231A/G429A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87L/T231A/L382M site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87L/T231A/L382M/G429A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87L/Y152F/T231A/L382M/G429A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B16 Y59W/Y87V/T231A site-directed mutagenesis Ruegeria sp. TM1040
2.6.1.B21 additional information mutational analysis of enzyme-substrate interactions and substrate specificity, overview. The evolved ATAs perform asymmetric synthesis of the respective R-amine with high conversions by using either alanine or isopropylamine Ruegeria sp. TM1040
2.6.1.B21 P281S site-directed mutagenesis, the mutation seems to have a negative effect on specific activity Ruegeria sp. TM1040
2.6.1.B21 Y152F site-directed mutagenesis, the mutation stabilizes the enzyme, the activity is slightly reduced compared to wild-type Ruegeria sp. TM1040
2.6.1.B21 Y59W/T231A site-directed mutagenesis, inactive mutant Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87F/T231A site-directed mutagenesis, inactive mutant Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87L/T231A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87L/T231A/G429A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87L/T231A/L382M site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87L/T231A/L382M/G429A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87L/Y152F/T231A/L382M/G429A site-directed mutagenesis, specific and enantioimeric conversion of 2,2-dimethyl-1-phenylpropan-1-amine compared to wild-type Ruegeria sp. TM1040
2.6.1.B21 Y59W/Y87V/T231A site-directed mutagenesis Ruegeria sp. TM1040

Organism

EC Number Organism UniProt Comment Textmining
2.6.1.B16 Ruegeria sp. TM1040
-
-
-
2.6.1.B21 Ruegeria sp. TM1040
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.6.1.B21 recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, dialysis, and ultrafiltration Ruegeria sp. TM1040

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.6.1.B16 0.0065
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87V/T231A Ruegeria sp. TM1040
2.6.1.B16 0.0115
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A Ruegeria sp. TM1040
2.6.1.B16 0.019
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/L382M Ruegeria sp. TM1040
2.6.1.B16 0.0324
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/P281S/G429A Ruegeria sp. TM1040
2.6.1.B16 0.0448
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/G429A Ruegeria sp. TM1040
2.6.1.B16 0.0771
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/L382M/G429A Ruegeria sp. TM1040
2.6.1.B21 0.0065
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87V/T231A Ruegeria sp. TM1040
2.6.1.B21 0.0115
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A Ruegeria sp. TM1040
2.6.1.B21 0.019
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/L382M Ruegeria sp. TM1040
2.6.1.B21 0.0324
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/P281S/G429A Ruegeria sp. TM1040
2.6.1.B21 0.0448
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/G429A Ruegeria sp. TM1040
2.6.1.B21 0.0771
-
 pH 8.0, 30°C, substrate rac-2,2-dimethyl-1-phenylpropan-1-amine, mutant Y59W/Y87L/T231A/L382M/G429A Ruegeria sp. TM1040

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.6.1.B16 2,2-dimethyl-1-phenylpropan-1-one + isopropylamine the conversion by all active enzyme mutants lays between 22 and 71% with isopropylamine as amine donor, with enantiomeric excess above 99%, low activity Ruegeria sp. TM1040 (R)-2,2-dimethyl-1-phenylpropan-1-amine + acetone
-
 r
2.6.1.B16 2,2-dimethyl-1-phenylpropan-1-one + L-alanine the conversion by all active enzyme mutants is complete with L-alanine as amine donor, with enantiomeric excess above 99% Ruegeria sp. TM1040 (R)-2,2-dimethyl-1-phenylpropan-1-amine + pyruvate
-
 r
2.6.1.B16 additional information engineered ATAs perform asymmetric synthesis of the respective R-amine with high conversions by using either alanine or isopropylamine as amine donor. Asymmetric synthesis of (R)-2,2-dimethyl-1-phenylpropan-1-amine by amino group transfer to 2,2-dimethyl-1-phenylpropan-1-one catalyzed by ATAs. Isopropylamine or alanine serve as the amine donors. Analysis of specific activities of Rsp-ATA mutant variants towards rac-amine 2,2-dimethyl-1-phenylpropan-1-amine. Enzyme-ligand interaction analysis, overview Ruegeria sp. TM1040 ?
-
-
2.6.1.B16 rac-2,2-dimethyl-1-phenylpropan-1-amine + pyruvate
-
 Ruegeria sp. TM1040 2,2-dimethyl-1-phenylpropan-1-one + L-alanine
-
 r
2.6.1.B21 2,2-dimethyl-1-phenyl-propan-1-one + isopropylamine the conversion by all active enzyme mutants lays between 22 and 71% with isopropylamine as amine donor, with enantiomeric excess above 99%, low activity Ruegeria sp. TM1040 (R)-2,2-dimethyl-1-phenylpropan-1-amine + acetone
-
 r
2.6.1.B21 2,2-dimethyl-1-phenyl-propan-1-one + L-alanine the conversion by all active enzyme mutants is complete with L-alanine as amine donor, with enantiomeric excess above 99% Ruegeria sp. TM1040 (R)-2,2-dimethyl-1-phenylpropan-1-amine + pyruvate
-
 r
2.6.1.B21 additional information engineered ATAs perform asymmetric synthesis of the respective R-amine with high conversions by using either alanine or isopropylamine as amine donor. Asymmetric synthesis of (R)-2,2-dimethyl-1-phenylpropan-1-amine by amino group transfer to 2,2-dimethyl-1-phenyl-propan-1-one catalyzed by ATAs. Isopropylamine or alanine serve as the amine donors. Analysis of specific activities of Rsp-ATA mutant variants towards rac-amine 2,2-dimethyl-1-phenylpropan-1-amine. Enzyme-ligand interaction analysis, overview Ruegeria sp. TM1040 ?
-
-
2.6.1.B21 rac-2,2-dimethyl-1-phenylpropan-1-amine + pyruvate
-
 Ruegeria sp. TM1040 2,2-dimethyl-1-phenyl-propan-1-one + L-alanine
-
 r

Synonyms

EC Number Synonyms Comment Organism
2.6.1.B16 amine transaminase
-
 Ruegeria sp. TM1040
2.6.1.B16 ATA
-
 Ruegeria sp. TM1040
2.6.1.B16 Rsp-ATA
-
 Ruegeria sp. TM1040
2.6.1.B21 ATA
-
 Ruegeria sp. TM1040
2.6.1.B21 Rsp-ATA
-
 Ruegeria sp. TM1040

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.6.1.B16 30
-
 assay at Ruegeria sp. TM1040
2.6.1.B21 30
-
 assay at Ruegeria sp. TM1040

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.6.1.B16 8 9 assay at Ruegeria sp. TM1040
2.6.1.B21 8 9 assay at Ruegeria sp. TM1040

Cofactor

EC Number Cofactor Comment Organism Structure
2.6.1.B16 pyridoxal 5'-phosphate PLP, dependent on Ruegeria sp. TM1040
2.6.1.B21 pyridoxal 5'-phosphate PLP, dependent on Ruegeria sp. TM1040

General Information

EC Number General Information Comment Organism
2.6.1.B16 evolution the ATA enzyme from Ruegeria sp. TM1040 (Rsp-ATA, PDB ID 3FCR) belongs to the ATAs of fold class I Ruegeria sp. TM1040
2.6.1.B21 evolution the ATA enzyme from Ruegeria sp. TM1040 (Rsp-ATA, PDB ID 3FCR) belongs to the ATAs of fold class I Ruegeria sp. TM1040