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Formation of Indigo by Recombinant Mammalian Cytochrome P450

Gillam, Elizabeth M. J. and Aguinaldo, Anna Marie A. and Notley, Lisa M. and Kim, Donghak and Mundkowski, Ralf G. and Volkov, Alexander A. and Arnold, Frances H. and Souček, Pavel and DeVoss, James J. and Guengerich, F. Peter (1999) Formation of Indigo by Recombinant Mammalian Cytochrome P450. Biochemical and Biophysical Research Communications, 265 (2). pp. 469-472. ISSN 0006-291X. https://resolver.caltech.edu/CaltechAUTHORS:20180823-144310509

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Abstract

The development of bicistronic systems for coexpression of recombinant human cytochrome P450 enzymes (P450s) with their redox partner, NADPH-cytochrome P450 reductase (NPR), has enabled P450 activity to be reconstituted within bacterial cells. During expression of recombinant P450 2E1 and some other forms, we observed the formation of a blue pigment in bacterial cultures. The pigment was extracted from cultures and shown to comigrate with standard indigo on TLC. UV-visible spectroscopy and mass spectrometric analysis provided further support for identification of the pigment as indigo. Indigo is known to form following the spontaneous oxidation of 3-hydroxyindole. Accordingly, we speculated that indole, formed as a breakdown product of tryptophan in bacteria, was hydroxylated by the P450 system, leading to indigo formation. Bacterial membranes containing recombinant P450 2E1 and human NPR were incubated in vitro with indole and shown to catalyze formation of a blue pigment in a time- and cofactor-dependent manner. These studies suggest potential applications of mammalian P450 enzymes in industrial indigo production or in the development of novel colorimetric assays based on indole hydroxylation.


Item Type:Article
Related URLs:
URLURL TypeDescription
https://doi.org/10.1006/bbrc.1999.1702DOIArticle
ORCID:
AuthorORCID
Arnold, Frances H.0000-0002-4027-364X
Additional Information:© 1999 Academic Press. Received 5 October 1999. This work was supported in part by National Health and Medical Research Council (Australia) Grant 951135 and Australian Research Council Grant A09937199 (E.M.J.G.), Maxygen Corp. (F.H.A.), Grant IGA 1850-5 of the Internal Grant Agency of the Czech Ministry of Health (P.S.), and Grants R35 CA44353 and P30 ES00267 from the National Institutes of Health (F.P.G.).
Funders:
Funding AgencyGrant Number
National Health and Medical Research Council (NHMRC)951135
Australian Research CouncilA09937199
Maxygen CorporationUNSPECIFIED
Czech Ministry of HealthIGA 1850-5
NIHR35 CA44353
NIHP30 ES00267
Issue or Number:2
Record Number:CaltechAUTHORS:20180823-144310509
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20180823-144310509
Official Citation:Elizabeth M.J. Gillam, Anna Marie A. Aguinaldo, Lisa M. Notley, Donghak Kim, Ralf G. Mundkowski, Alexander A. Volkov, Frances H. Arnold, Pavel Souček, James J. DeVoss, F.Peter Guengerich, Formation of Indigo by Recombinant Mammalian Cytochrome P450, Biochemical and Biophysical Research Communications, Volume 265, Issue 2, 1999, Pages 469-472, ISSN 0006-291X, https://doi.org/10.1006/bbrc.1999.1702. (http://www.sciencedirect.com/science/article/pii/S0006291X99917023)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:89100
Collection:CaltechAUTHORS
Deposited By: George Porter
Deposited On:23 Aug 2018 22:22
Last Modified:03 Oct 2019 20:13

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