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Precision Molecular Editing: Predicting Substrate Scope and Regiochemistry for CHEESY1, a Flavin Dependent
Ying Zhang1,2, Olena Holodaieva1,2, Yunpeng Wang1,2
1EaStCHEM School of Chemistry, University of St Andrews, North Haugh, St Andrews KY16 9ST, Fife, U.K.
ACS Catalysis
|November 27, 2025
Summary
Scientists discovered a new enzyme, CHEESY1, from cheese microbes that precisely activates C-H bonds in medicinally relevant molecules. This enzyme enables targeted halogenation, revolutionizing chemical synthesis and molecule making.
Area of Science:
- Biocatalysis and Synthetic Chemistry
- Enzymology and Microbial Discovery
Background:
- C-H activation is crucial for molecule synthesis, but selective functionalization of heteroaromatic compounds remains challenging.
- Halogenases are enzymes that catalyze halogenation, with many known from marine environments, but their presence and utility in other saline ecosystems are less explored.
- Existing halogenase discovery is often linked to known natural product biosynthetic gene clusters, limiting the exploration of novel enzymatic functions.
Purpose of the Study:
- To discover and characterize novel halogenase enzymes from non-traditional saline environments, specifically fermented foods.
- To develop a predictive in silico approach for identifying halogenase function and substrate scope independent of known biosynthetic pathways.
- To demonstrate the laboratory validation and synthetic utility of a newly discovered halogenase for precision C-H activation and halogenation.
Main Methods:
- Genome mining and in silico analysis to identify potential halogenase genes in microbes from brined cheese.
- Development of computational methods to predict enzyme activity, substrate specificity, and regiochemistry for non-native substrates.
- Laboratory-based experimental validation of the in silico predictions using the discovered halogenase (CHEESY1).
Main Results:
- Discovery and characterization of CHEESY1 (Chemistry Helper Enzyme Enabling SelectivitY1), a novel halogenase from a cheese-associated microbe.
- Demonstration of CHEESY1's ability to regioselectively halogenate a diverse range of medicinally relevant heterocycles, including quinolines, isoquinoline, phenylpyrazole, and flavonoids.
- Validation of the in silico prediction approach for halogenase discovery and substrate scope determination.
Conclusions:
- The discovery of CHEESY1 expands the known diversity of halogenases to fermented food environments.
- The predictive in silico methodology enables efficient discovery and functional characterization of novel halogenases and their substrates.
- CHEESY1 represents a powerful new tool for precision C-H activation and halogenation, offering significant potential for advancing chemical synthesis and drug discovery.
