Members of the cytochrome P450 monooxygenase family CYP268 are found across a broad range of Mycobacterium species including the pathogens Mycobacterium avium, M. colombiense, M. kansasii, and M. marinum. CYP268A2, from M. marinum, which is the first member of this family to be studied, was purified and characterised. CYP268A2 was found to bind a variety of substrates with high affinity, including branched and straight chain fatty acids (C10–C12), acetate esters, and aromatic compounds. The enzyme was also found to bind phenylimidazole inhibitors but not larger azoles, such as ketoconazole. The monooxygenase activity of CYP268A2 was efficiently reconstituted using heterologous electron transfer partner proteins. CYP268A2 hydroxylated geranyl acetate and trans-pseudoionone at a terminal methyl group to yield (2E,6E)-8-hydroxy-3,7-dimethylocta-2,6-dien-1-yl acetate and (3E,5E,9E)-11-hydroxy-6,10-dimethylundeca-3,5,9-trien-2-one, respectively. The X-ray crystal structure of CYP268A2 was solved to a resolution of 2.0 Å with trans-pseudoionone bound in the active site. The overall structure was similar to that of the related phytanic acid monooxygenase CYP124A1 enzyme from Mycobacterium tuberculosis, which shares 41% sequence identity. The active site is predominantly hydrophobic, but includes the Ser99 and Gln209 residues which form hydrogen bonds with the terminal carbonyl group of the pseudoionone. The structure provided an explanation on why CYP268A2 shows a preference for shorter substrates over the longer chain fatty acids which bind to CYP124A1 and the selective nature of the catalysed monooxygenase activity.
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February 2018
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The X-ray crystal structure of the substrate binding pocket of cytochrome P450 enzyme CYP268A2, from Mycobacterium marinum, with the molecule pseudoionone bound in the active site. In this issue of the Biochemical Journal, Bell et al. show how the substrate becomes completely enclosed by the active site of the enzyme; for details see pages 705–722.
Research Article|
February 16 2018
Structural and functional characterisation of the cytochrome P450 enzyme CYP268A2 from Mycobacterium marinum
Stella A. Child;
Stella A. Child
1Department of Chemistry, School of Physical Sciences, University of Adelaide, Adelaide, SA 5005, Australia
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Elise F. Naumann;
Elise F. Naumann
1Department of Chemistry, School of Physical Sciences, University of Adelaide, Adelaide, SA 5005, Australia
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John B. Bruning;
John B. Bruning
2School of Biological Sciences, University of Adelaide, Adelaide, SA 5005, Australia
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Stephen G. Bell
1Department of Chemistry, School of Physical Sciences, University of Adelaide, Adelaide, SA 5005, Australia
Correspondence: Stephen G. Bell (stephen.bell@adelaide.edu.au)
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Publisher: Portland Press Ltd
Received:
December 13 2017
Revision Received:
January 10 2018
Accepted:
January 16 2018
Accepted Manuscript online:
January 17 2018
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 2018 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society
2018
Biochem J (2018) 475 (4): 705–722.
Article history
Received:
December 13 2017
Revision Received:
January 10 2018
Accepted:
January 16 2018
Accepted Manuscript online:
January 17 2018
Citation
Stella A. Child, Elise F. Naumann, John B. Bruning, Stephen G. Bell; Structural and functional characterisation of the cytochrome P450 enzyme CYP268A2 from Mycobacterium marinum. Biochem J 28 February 2018; 475 (4): 705–722. doi: https://doi.org/10.1042/BCJ20170946
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