Reaktion #658765

Reaktionsgleichung

pyridoxal phosphate

ampicillin

(S)-7-methoxy-2-aminotetralin

potassium phosphate

→

7-Methoxy-2-Tetralone

Lösungsmittel

hydrochloric acid

Reaktionsbedingungen

Aufarbeitung

1. 1
   Sonstigecoli obtained in Example 2
2. 2
   Sonstigethe mixture was sonicated so that
3. 3
   Extraktiona cell-free extract
4. 4
   Sonstigewas obtained
5. 5
   ExtraktionSubsequently, 100 μL of the cell-free extract
6. 6
   workup.ADDITIONwas added to 305 μL of a substrate solution
7. 7
   Sonstigeto react at 30° C. for 1 hour
8. 8
   Sonstigethe reaction

Vorschrift

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.

Quelle