Construction and Optimization of a Heterologous Pathway for Protocatechuate Catabolism in Escherichia coli Enables Bioconversion of Model Aromatic Compounds
[2];
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- Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division
- Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Chemical Sciences Division
- Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center
- Michigan State Univ., East Lansing, MI (United States)
The production of biofuels from lignocellulose yields a substantial lignin by-product stream that currently has few applications. Biological conversion of lignin-derived compounds into chemicals and fuels has the potential to improve the economics of lignocellulose-derived biofuels, but few microbes are able both to catabolize lignin-derived aromatic compounds and to generate valuable products. WhileEscherichia colihas been engineered to produce a variety of fuels and chemicals, it is incapable of catabolizing most aromatic compounds. Therefore, we engineeredE. colito catabolize protocatechuate, a common intermediate in lignin degradation, as the sole source of carbon and energy via heterologous expression of a nine-gene pathway fromPseudomonas putidaKT2440. Then, we used experimental evolution to select for mutations that increased growth with protocatechuate more than 2-fold. Increasing the strength of a single ribosome binding site in the heterologous pathway was sufficient to recapitulate the increased growth. After optimization of the core pathway, we extended the pathway to enable catabolism of a second model compound, 4-hydroxybenzoate. These engineered strains will be useful platforms to discover, characterize, and optimize pathways for conversions of lignin-derived aromatics. IMPORTANCELignin is a challenging substrate for microbial catabolism due to its polymeric and heterogeneous chemical structure. Therefore, engineering microbes for improved catabolism of lignin-derived aromatic compounds will require the assembly of an entire network of catabolic reactions, including pathways from genetically intractable strains. By constructing defined pathways for aromatic compound degradation in a model host would allow rapid identification, characterization, and optimization of novel pathways. Finally, we constructed and optimized one such pathway inE. colito enable catabolism of a model aromatic compound, protocatechuate, and then extended the pathway to a related compound, 4-hydroxybenzoate. This optimized strain can now be used as the basis for the characterization of novel pathways.
- Research Organization:
- Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
- Sponsoring Organization:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- DOE Contract Number:
- AC05-00OR22725; AC02-05CH1231
- OSTI ID:
- 1394233
- Journal Information:
- Applied and Environmental Microbiology, Vol. 83, Issue 18; ISSN 0099-2240
- Publisher:
- American Society for Microbiology
- Country of Publication:
- United States
- Language:
- English
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