Reaktionsgleichung
[O:1]=C(CCC(O)=O)C(O)=O.O=C1O[C@H]([C@H](CO)O)C(O)=C1O.C1N(CCO)CCN(CCS(O)(=O)=O)C1.[NH2:38][C@H:39]([C:44]([OH:46])=[O:45])[C@H:40]([CH2:42][CH3:43])[CH3:41].N[C@H](C(O)=O)C(C)C.N[C@H](C(O)=O)CCC(O)=O.N[C@H](C(O)=O)CCCCN>>[OH:1][CH:42]([CH3:43])[C@H:40]([CH3:41])[C@@H:39]([C:44]([OH:46])=[O:45])[NH2:38]Komponenten
Versuchsvorschrift
By using a cell lysate prepared from 2-e-2 cells at OD660 of 7 according to the method described in <5>, the reaction characteristics for various amino acids were evaluated. The cell lysate and a substrate solution were mixed, then the reaction proceeded at 30° C. for 1 hour, and the production of new substances was evaluated by TLC or amino acid analysis. The substrate reaction mixture contained 5 mM amino acid, 5 mM Fe2+, 5 mM 2-oxoglutaric acid, 5 mM ascorbic acid, and 100 mM HEPES (pH 7). In addition to L-isoleucine, the amino acids L-leucine, L-valine, L-glutamic acid, and L-lysine were each individually evaluated. The 4-hydroxyisoleucine which was produced was analyzed by the method described in Example 1. The results are shown in Table 6. Production of amino acids other than L-isoleucine was not observed. Therefore, it was suggested that this enzyme was an isoleucine-specific dioxygenase.
Aufarbeitung
- additionThe cell lysate and a substrate solution were mixed
- customThe substrate reaction mixture
Quellen und Nachweise
- ORD-Eintrag
- ord-eb25dca2d1d14ea28fefb8291ea774c1
- Quelldatensatz
- uspto-grants-2013_02
- Patentnummer
- US08367382B2
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