Isopropanol → NADPH

HRID 2400244Notes

Reaction equation

HRID 2400244 reaction equation
C(O)(C)C.[CH:5]1[CH:10]=[N+:9]([C@@H:11]2[O:15][C@H:14]([CH2:16][O:17][P:18]([O:21][P:22]([O:25][CH2:26][C@H:27]3[O:31][C@@H:30]([N:32]4[C:36]5[N:37]=[CH:38][N:39]=[C:40]([NH2:41])[C:35]=5[N:34]=[CH:33]4)[C@H:29]([O:42][P:43]([OH:46])([OH:45])=[O:44])[C@@H:28]3[OH:47])([OH:24])=[O:23])([OH:20])=[O:19])[C@@H:13]([OH:48])[C@H:12]2[OH:49])[CH:8]=[C:7]([C:50]([NH2:52])=[O:51])[CH:6]=1>CC(C)=O>[CH:38]1[N:39]=[C:40]([NH2:41])[C:35]2[N:34]=[CH:33][N:32]([C@@H:30]3[O:31][C@H:27]([CH2:26][O:25][P:22]([O:21][P:18]([O:17][CH2:16][C@H:14]4[O:15][C@@H:11]([N:9]5[CH:8]=[C:7]([C:50]([NH2:52])=[O:51])[CH2:6][CH:5]=[CH:10]5)[C@H:12]([OH:49])[C@@H:13]4[OH:48])([OH:20])=[O:19])([OH:24])=[O:23])[C@@H:28]([OH:47])[C@H:29]3[O:42][P:43]([OH:46])([OH:45])=[O:44])[C:36]=2[N:37]=1

Participants

4

Reactant 2

HCID 3776IsopropanolHCID 5886NADP nicotinamide-adenine-dinucleotide phosphate

Product 1

HCID 5884NADPH

Reagents, catalysts & solvents 1

HCID 180AcetoneSolvent

Procedure

This example illustrates a prescreening assay used to identify variant genes encoding ketoreductases capable of reducing isopropanol in the presence of NADP+ to produce acetone and NADPH. An E. coli colony containing a plasmid encoding an engineered ketoreductase was picked using a Q-Bot® robotic colony picker (Genetix USA, Inc., Boston, Mass.) into 96-well shallow well microtiter plates containing 180 μL Terrific Broth (TB), 1% glucose and 30 μg/mL chloramphenicol (CAM). Cells were grown overnight at 30° C. with shaking at 200 rpm. A 10 μL aliquot of this culture was then transferred into 96-deep well plates containing 390 μL Terrific Broth (TB), 1 mM MgSO4 and 30 μg/mL CAM. After incubation of the deep-well plates at 30° C. with shaking at 250 rpm for 2 to 3 hours, recombinant gene expression within the cultured cells was induced by addition of IPTG to a final concentration of 1 mM. The plates were then incubated at 30° C. with shaking at 250 rpm for 18 hours.

Sources & evidence

ORD record
ord-f18ad8b743674e01a71261e3664ee9be
Source dataset
uspto-grants-2014_08
Patent number
US08796002B2

Reaction notes

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