Optimized Spatial Configuration of Heterogeneous Biocatalysts Maximizes Cell-Free Biosynthesis of ω‑Hydroxy and ω‑Amino Acids

Cell-free biocatalysis is gaining momentum in producing value-added chemicals, particularly in stepwise reaction cascades. However, the stability of enzyme cascades in industrial settings is often compromised when free enzymes are involved. In this study, we have developed a stable multifunctional h...

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Published in:ACS sustainable chemistry & engineering 2024-06, Vol.12 (25), p.9474-9489
Main Authors: Santiago-Arcos, Javier, Velasco-Lozano, Susana, Diamanti, Eleftheria, Benítez-Mateos, Ana I., Grajales-Hernández, Daniel, Paradisi, Francesca, López-Gallego, Fernando
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Language:English
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Summary:Cell-free biocatalysis is gaining momentum in producing value-added chemicals, particularly in stepwise reaction cascades. However, the stability of enzyme cascades in industrial settings is often compromised when free enzymes are involved. In this study, we have developed a stable multifunctional heterogeneous biocatalyst coimmobilizing five enzymes on microparticles to transform 1,ω-diols into 1,ω-hydroxy acids. We improved the operational efficiency and stability of the heterogeneous biocatalyst by fine-tuning the enzyme loading and spatial organization. Stability issues are overcome through postimmobilization polymer coating. The general applicability of this heterogeneous biocatalyst is demonstrated by its scale-up in both batch and packed bed reactors, allowing a product yield of >80%. The continuous process is fed with H2O2 as the oxygen source, reaching a space-time yield (STY) of 0.76 g·L–1·h–1, maintained for the first 12 h. Finally, this flow system is telescoped with a second plug-flow reactor packed with a different heterogeneous biocatalyst integrating an additional transaminase. As a result, this 6-enzyme 2-reactor system sequentially transforms 1,ω-diols into 1,ω-amino acids while in situ recycling NAD+, depleting H2O2, and generating O2.
ISSN:2168-0485
2168-0485
DOI:10.1021/acssuschemeng.4c02396