Formation of indigo by recombinant mammalian cytochrome P450.

Gillam EM, Aguinaldo AM, Notley LM, Kim D, Mundkowski RG, Volkov AA, Arnold FH, Soucek P, DeVoss JJ, Guengerich FP
Biochem Biophys Res Commun. 1999 265 (2): 469-72

PMID: 10558891 · DOI:10.1006/bbrc.1999.1702

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.

Copyright 1999 Academic Press.

MeSH Terms (11)

Cytochrome P-450 CYP2E1 Cytochrome P-450 Enzyme System Escherichia coli Genes Humans Indigo Carmine Indoles In Vitro Techniques NADPH-Ferrihemoprotein Reductase Pigments, Biological Recombinant Proteins

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