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Updated: May 16, 2026

Fluorescence-Based Detection of FEN1 Nuclease Activity and Screening of Small-Molecule Inhibitors
Published on: June 27, 2025
Catalytic control of enzymatic fluorine specificity
Amy M Weeks1, Michelle C Y Chang
1Department of Chemistry, University of California, Berkeley, CA 94720, USA.
Abstract:
The investigation of unique chemical phenotypes has led to the discovery of enzymes with interesting behaviors that allow us to explore unusual function. The organofluorine-producing microbe Streptomyces cattleya has evolved a fluoroacetyl-CoA thioesterase (FlK) that demonstrates a surprisingly high level of discrimination for a single fluorine substituent on its substrate compared with the cellularly abundant hydrogen analog, acetyl-CoA. In this report, we show that the high selectivity of FlK is achieved through catalysis rather than molecular recognition, where deprotonation at the C(α) position to form a putative ketene intermediate only occurs on the fluorinated substrate, thereby accelerating the rate of hydrolysis 10(4)-fold compared with the nonfluorinated congener. These studies provide insight into mechanisms of catalytic selectivity in a native system where the existence of two reaction pathways determines substrate rather than product selection.
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