Identification of broad specificity P450CAM variants by primary screening against indole as substrate

Chemical Communications

Volumn , Issue 29, 2005, Pages 3652-3654

  Çelik, Ayhan ^a   Speight, Robert E ^a   Turner, Nicholas J ^a  

^a UNIVERSITY OF EDINBURGH   (United Kingdom)

Author keywords

[No Author keywords available]

Indexed keywords

CYTOCHROME P450; INDOLE DERIVATIVE; INDOXYL; UNCLASSIFIED DRUG;

ARTICLE; CATALYSIS; ENZYME ACTIVITY; ENZYME ANALYSIS; ENZYME SPECIFICITY; ENZYME STRUCTURE; GAS CHROMATOGRAPHY; HYDROXYLATION; MASS SPECTROMETRY; OXIDATION; SCREENING;

EID: 23644456287     PISSN: 13597345     EISSN: None     Source Type: Journal    
DOI: 10.1039/b506156c     Document Type: Article

References (14)

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11. CAM proteins which would be used for the primary screening experiment against new substrates (i.e. indole).
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14. 600) at 30 °C. Induction was achieved using 0.02% of L-arabinose. After 30 minutes of induction, indole (1 mM final concentration) was delivered in DMSO (0.5%) into the medium. After 24 hours biotransformation at 30 °C, cells were harvested at 10 K rpm for 30 minutes. Cells then were washed with water. The residual blue pellet was extracted with tetrahydrofuran (THF, 3 × 25 ml). The solvent was evaporated to dryness and red pigment extracted from the residue with ethanol (5 × 10 ml). The ethanol extract was evaporated to dryness and a silica gel column used for separation and purification. (DC 60 Merck, eluted with THF: petroleum ether 1:2).