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. 2021 Mar 5;11:5290. doi: 10.1038/s41598-021-84859-6

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© The Author(s) 2021

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 licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence 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 licence, visit http://creativecommons.org/licenses/by/4.0/.

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The OsmY-Tfu0937 activity in the spent medium of single site-directed mutants was measured with the pNPG assay using an absorbance at 405 nm. Cells were cultivated in 20-mL of 2 × TY auto-induction media at 37 °C. Culture was inoculated using 1:100 dilution and 0.5 mL culture was sampled at various time points for activity assay. Experiments were performed in triplicate. The samples are represented using different colours; OsmY(WT)—black line, OsmY(L6P)—blue line, OsmY(S154R)—orange line and OsmY(V191E)—red line.