Pyridoxal 5'-phosphate → 7-Methoxy-2-tetralone
Reaction equation
CC1(C)S[C@@H]2[C@H](NC([C@H](N)C3C=CC=CC=3)=O)C(=[O:9])N2[C@H]1C(O)=O.P([O-])([O-])([O-])=O.[K+].[K+].[K+].CC1N=CC(COP(O)(O)=O)=C(C=O)C=1O.[CH3:49][O:50][C:51]1[CH:60]=[C:59]2[C:54]([CH2:55][CH2:56][C@H:57](N)[CH2:58]2)=[CH:53][CH:52]=1>Cl>[CH3:49][O:50][C:51]1[CH:60]=[C:59]2[C:54]([CH2:55][CH2:56][C:57](=[O:9])[CH2:58]2)=[CH:53][CH:52]=1Participants
Reactant 4
Product 1
Reagents, catalysts & solvents 1
Conditions
- Temperature
- 30 °C
Procedure
The recombinant E. coli obtained in Example 2 was inoculated into 5 mL of a 2× YT medium containing 200 μg/mL of ampicillin and cultured with shaking at 30° C. for 28 hours. To 500 μL of the resulting culture liquid was added 500 μL of a 0.2 M potassium phosphate buffer (pH 7.5) containing 1 mM pyridoxal phosphate, and the mixture was sonicated so that a cell-free extract was obtained. Subsequently, 100 μL of the cell-free extract was added to 305 μL of a substrate solution having the composition shown below. After the mixture was allowed to react at 30° C. for 1 hour, 50 μL of 6 N hydrochloric acid was added to stop the reaction. The optical purity of the produced (S)-7-methoxy-2-aminotetralin was measured using HPLC under the conditions shown below. As a result, the modified enzymes shown in Table 2 had stereoselectivity higher than that of the wild-type.
Workup
- customcoli obtained in Example 2
- customthe mixture was sonicated so that
- extractiona cell-free extract
- customwas obtained
- extractionSubsequently, 100 μL of the cell-free extract
- additionwas added to 305 μL of a substrate solution
- customto react at 30° C. for 1 hour
- customthe reaction
Sources & evidence
- ORD record
- ord-8f3a57902add4c02a3519cbb261a05ab
- Source dataset
- uspto-grants-2015_05
- Patent number
- US09029113B2
Reaction notes
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