Biocatalytic control of site-selectivity and chain length selectivity in radical amino acid halogenases

Biocatalytic C???H activation has the potential to merge enzymatic and synthetic strategies for bond formation. FeII/??KG-dependent halogenases are particularly dis-tinguished for their ability both to control selective C???H activation as well as to direct group transfer of a bound anion along a reaction axis separate from oxygen rebound, enabling the development of new transformations. In this context, we elucidate the basis for the selectivity of enzymes that perform selective halogena-tion to yield 4-Cl-lysine (BesD), 5-Cl-lysine (HalB), and 4-Cl-ornithine (HalD), allowing us to probe how site-selectivity and chain length selectivity are achieved. We now report the crystal structure of the HalB and HalD, revealing the key role of the substrate-binding lid in positioning the substrate for C4 vs C5 chlorination and recognition of lysine vs ornithine. Targeted engineering of the substrate-binding lid further demonstrates that these selectivities can be altered or switched, showcasing the potential to develop halogenases for biocatalytic applications.

Automated summary

This study investigates the application of enzyme-catalyzed C-H activation in selective halogenation, and elucidates the crucial role of substrate-binding lid in selective halogenation through crystal structures of HalB and HalD, providing guidance for the development of halogenases with biocatalytic applications.