Abstract
Aryl-O-demethylation is a common rate-limiting step in the catabolism of lignin-related compounds, including guaiacol. Here we used randomly barcoded transposon insertion sequencing (RB-TnSeq) in the bacterium Novosphingobium aromaticivorans to identify a Rieske-type guaiacol O-demethylase, GdmA. Similarity searches identified GdmA homologs in other bacteria, along with candidate reductase partners, denoted GdmB. GdmAB combinations were biochemically characterized for activity with several lignin-related substrates. Structural and sequence comparisons of vanillate- and guaiacol-specific O-demethylase active sites revealed conserved hallmarks of substrate specificity. GdmAB combinations were also evaluated in Pseudomonas putida KT2440, which does not natively utilize guaiacol. GdmAB from Cupriavidus necator N-1 demonstrated the highest rate of guaiacol turnover in vitro and in engineered P. putida strains and notably higher catalytic efficiency than a cytochrome P450 system (GcoAB) and the vanillate Rieske-type O-demethylase from P. putida (VanAB). The GdmAB O-demethylases described here expand the suite of options for microbial conversion of a model lignin-derived substrate.
Original language | American English |
---|---|
Pages (from-to) | 1989-2011 |
Number of pages | 23 |
Journal | Chem Catalysis |
Volume | 2 |
Issue number | 8 |
DOIs | |
State | Published - 18 Aug 2022 |
Bibliographical note
Publisher Copyright:© 2022 Elsevier Inc.
NREL Publication Number
- NREL/JA-2800-81928
Keywords
- biocatalysis
- biological funneling
- Cupriavidus necator
- microbial lignin conversion
- Novosphingobium aromaticivorans
- O-demethylation
- Pseudomonas putida KT2440
- Rieske non-heme iron monooxygenase
- SDG 7: Affordable and clean energy
- SDG 9: Industry, innovation, and infrastructure
- SDG13: Climate action
- Sphingomonas wittichii