Changing the electron donor improves azoreductase dye degrading activity at neutral pH

Jingxian Qi, Caroline E. Paul, Frank Hollmann, Dirk Tischler

Research output: Contribution to journalArticleScientificpeer-review

16 Citations (Scopus)
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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.

Original languageEnglish
Pages (from-to)17-19
Number of pages3
JournalEnzyme and Microbial Technology: biotechnology research and reviews
Volume100
DOIs
Publication statusPublished - 2017

Keywords

  • 1-benzyl-1,4-dihydronicotinamide
  • Azo dyes
  • Azoreductase
  • Methyl red degradation
  • Nicotinamide cofactor mimics
  • Rhodococcus

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