Screening for cytochrome P450 reactivity by harnessing catalase as reporter enzyme

  • Rabe K
  • Spengler M
  • Erkelenz M
 et al. 
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Abstract

Cytochrome P450 enzymes are known to catalyze a variety of reactions that are difficult to perform by standard organic synthesis, such as the oxidation of unactivated C--C bonds. Cytochrome P450 enzymes can also be used in artificial systems in which organic peroxides act as cosubstrates. To find substrates that are converted by a certain P450 catalyst in the presence of an organic peroxide, various screening assays have been established, however, most of them are limited to one or only a few specific substrates. Here, we report a simple and rapid screening assay that works independently of the nature of the substrate and utilizes a previously undescribed reactivity of catalase as reporter enzyme. In an initial demonstration of this assay, we screened 180 enzyme/peroxide/substrate combinations for potential bioconversions. As shown by subsequent verification of the screening results with liquid chromatography/multistage mass spectrometry (LC/MS(n)), we were able to identify three new substrates for the enzyme CYP152A1 and at least two previously undescribed conversions by the enzyme CYP119.

Author-supplied keywords

  • Biocatalysis
  • Cytochrome P450
  • Enzymes
  • Highthroughput screening
  • Oxidases

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Authors

  • Kersten S. Rabe

  • Mark Spengler

  • Michael Erkelenz

  • Joachim Müller

  • Valerie J. Gandubert

  • Heiko Hayen

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