A High-Throughput Mass Spectrometric Enzyme Activity Assay Enabling the Discovery of Cytochrome P450 Biocatalysts

Research output: Contribution to journalJournal article – Annual report year: 2019Researchpeer-review


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Assaying for enzymatic activity is a persistent bottleneck in biocatalyst and drug development. Existing high-throughput assays for enzyme activity tend to be applicable only to a narrow range of biochemical transformations, whereas universal enzyme characterization methods usually require chromatography to determine substrate turnover, greatly diminishing throughput. We present an enzyme activity assay that allows the high-throughput mass-spectrometric detection of enzyme activity in complex matrices without the need for a chromatographic step. This technology, which we call probing enzymes with click-assisted NIMS (PECAN), can detect the activity of medically and biocatalytically significant cytochrome P450s in cell lysate, microsomes, and bacteria. Using this approach, a cytochrome P450BM3 mutant library was successfully screened for the ability to catalyze the oxidation of the sesquiterpene valencene.
Original languageEnglish
Issue number30
Pages (from-to)10114-10119
Number of pages7
Publication statusPublished - 2019
CitationsWeb of Science® Times Cited: No match on DOI

    Research areas

  • Biocatalysis, Cytochrome P450, Enzyme assays, High-throughput screening, Mass spectrometry

ID: 182567106