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. 2019 Oct 29;9:15479. doi: 10.1038/s41598-019-52077-w

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Rescreening of variants from MP-CE based selection, (a,c), and NADPH depletion based selection (b,d). Identified P450 BM3 variants from both screening methods were measured in triplicates and compared to the P450 BM3 WT (grey column) and EV (negative control). Variants which showed an increase of factor ≥1.3 (2.5-fold standard deviation, dashed line) in 4-hydroxy-isophorone formation (dashed columns) within the CE based selection (a) or NADPH oxidation rate (white columns) within the NADPH depletion based selection (b) were selected for further characterization. For a full picture, variants from the CE based selection (c) and NADPH depletion based selection (d) were screened with the opposite platform, however, results not considered for selection. (measurements: n = 4, mean ± s.e.; in biologically independent experiments) Substitutions of variants see Supplementary Table S2.