Print Email Facebook Twitter Changing the electron donor improves azoreductase dye degrading activity at neutral pH Title Changing the electron donor improves azoreductase dye degrading activity at neutral pH Author Qi, Jingxian (University of Technology Bergakademie Freiberg) Paul, C.E. (TU Delft BT/Biocatalysis) Hollmann, F. (TU Delft BT/Biocatalysis) Tischler, Dirk (University of Technology Bergakademie Freiberg) Date 2017 Abstract The oxygen-insensitive azoreductase AzoRo originating from Rhodococcus opacus 1CP was found to be most active at low pH (ca. 4) and high temperature (ca. 50 °C). AzoRo is not an efficient biocatalyst when used at low pH due to stability problems. To overcome this issue, we discovered that AzoRo accepts an alternative electron donor, 1-benzyl-1,4-dihydronicotinamide (BNAH), which allows fast turnover at neutral pH. In order to screen this nicotinamide coenzyme mimic as a source of electrons, AzoRo-catalysed reactions were run under neutral conditions, under which typically slow rates are observed with NADH. For the reduction of 1 azo bond by azoreductases 2 mol nicotinamide coenzyme are needed. AzoRo displayed Methyl Red (MR) reduction activities with NADH and NADPH of 5.49 ± 0.14 U mg−1 and 4.96 ± 0.25 U mg−1, respectively, whereas with BNAH it displayed 17.01 ± 0.74 U mg−1 (following BNAH oxidation) and 7.16 ± 0.06 U mg−1 (following MR reduction). Binding of BNAH to AzoRo was determined with a Km of 18.75 ± 2.45 μM (BNAH oxidation) and 12.45 ± 0.47 μM (MR reduction). In order to show applicability of this system an upscaled reaction was performed using 78.6 μg of purified AzoRo to convert 2.96 μmol of MR (total reaction volume: 40 ml) within a 1 h reaction. Subject 1-benzyl-1,4-dihydronicotinamideAzo dyesAzoreductaseMethyl red degradationNicotinamide cofactor mimicsRhodococcus To reference this document use: http://resolver.tudelft.nl/uuid:023e0c18-7509-4709-b2bf-e5edd58c2fe5 DOI https://doi.org/10.1016/j.enzmictec.2017.02.003 Embargo date 2018-02-16 ISSN 0141-0229 Source Enzyme and Microbial Technology: biotechnology research and reviews, 100, 17-19 Bibliographical note Accepted Author Manuscript Part of collection Institutional Repository Document type journal article Rights © 2017 Jingxian Qi, C.E. Paul, F. Hollmann, Dirk Tischler Files PDF manuscript_rev_DT.pdf 1.02 MB Close viewer /islandora/object/uuid:023e0c18-7509-4709-b2bf-e5edd58c2fe5/datastream/OBJ/view