Engineering CYP153A(M.aq) to Oxyfunctionalize its Inhibitor Dodecylamine Using a LC/MS Based Rapid Flow Analysis Screening

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Authors

RAPP Lea R. MARQUES Sérgio Manuel NEBEL Bernd DAMBORSKÝ Jiří HAUER Bernhard

Year of publication 2022
Type Article in Periodical
Magazine / Source ChemCatChem
MU Faculty or unit

Faculty of Science

Citation
Web https://chemistry-europe.onlinelibrary.wiley.com/doi/10.1002/cctc.202101648
Doi http://dx.doi.org/10.1002/cctc.202101648
Keywords biocatalysis; cytochrome P450; docking; enzyme engineering; molecular dynamics
Description The catalytic space of the P450 monooxygenase CYP153A(M.aq) was opened from a terminal (omega-) fatty acid hydroxylase to a catalyst capable of performing omega-hydroxylation of dodecylamine, which is a potent inhibitor for the wild-type enzyme. A simple screening method named Rapid-flow Analysis of Product Peaks (RAPP) was established and applied to measure saturation libraries directly from a 96-deepwell plate in 36 seconds per sample. The obtained variants are less inhibited by the amine, although concurrently show less affinity towards the acid. Molecular modelling and molecular dynamics simulations showed significant effects of the mutations on the substrate tunnel architectures.
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