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Versatile CO2 Transformations into Complex Products: A One-pot Two-step Strategy
Published on: November 9, 2019
Enzyme-Directed Charge Redistribution Controls Substrate Regioselectivity in CO2 Fixation by Salicylic Acid
Xianglong Li1, Zhicong He2, Haoxian Huang1
1MOE Key Laboratory of Bio-Intelligent Manufacturing, School of Bioengineering, Dalian University of Technology, Dalian, Liaoning 116024, China.
Journal of Agricultural and Food Chemistry
|June 29, 2026
Summary
Enzyme fields alter substrate charge distribution, enhancing regioselectivity in carboxylation reactions. This study reveals a novel mechanism for improving enzyme activity and selectivity in chemical synthesis.
Area of Science:
- Biocatalysis
- Enzyme Engineering
- Agricultural Chemistry
Background:
- 2,4-Dihydroxybenzoic acid is a key intermediate in agricultural and food chemistry.
- Salicylic acid decarboxylase (SAD) catalyzes resorcinol carboxylation but suffers from low regioselectivity and activity.
- Understanding enzyme-induced substrate modification is crucial for improving biocatalytic processes.
Purpose of the Study:
- To investigate how the enzyme's microenvironment (enzyme field) influences resorcinol's charge distribution and regioselectivity.
- To elucidate the mechanism of SAD-catalyzed carboxylation.
- To engineer a more selective and active SAD variant.
Main Methods:
- Computational calculation of charge distributions under various enzyme fields.
- Enzymatic catalytic assays to correlate regioselectivity with carbon atom electronegativity.
- Site-directed mutagenesis to create enzyme variants (e.g., K23A/Y64T/E291D/Y27A).
- QM/MM metadynamics simulations to propose the catalytic mechanism.
Main Results:
- Enzyme fields induce charge redistribution in resorcinol, altering regioselectivity.
- A strong correlation was observed between regioselectivity and the electronegativity of specific carbon atoms.
- The engineered variant K23A/Y64T/E291D/Y27A achieved 99% C4-carboxylation selectivity (up from 20%) and a 69-fold increase in kcat/Km.
- A mechanism involving HCO3-mediated proton transfer was proposed.
Conclusions:
- Enzyme-induced substrate charge distribution can be repurposed to control regioselectivity.
- The study provides a mechanistic understanding of SAD-catalyzed resorcinol carboxylation.
- Enzyme engineering strategies based on understanding the enzyme field can significantly enhance biocatalytic efficiency and selectivity.
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