Structure‐Guided Mutagenesis Reveals the Catalytic Residue that Controls the Regiospecificity of C6‐Indole Prenyltransferases

The functionalization of indole-derived compounds modifies their physicochemical properties and/or pharmacological activities. Activation of the indole C6 position is challenging. Yet, certain indole prenyltransferases (IPTs) catalyze this reaction efficiently, regiospecifically and under physiological conditions. Through enzyme engineering, in vitro enzymatic reactions and NMR experiments, guided by protein structure analyses, the mechanism was verified by which C6 IPTs catalyze their reactions.

Abstract

Indole is a significant structural moiety and functionalization of the C−H bond in indole-containing molecules expands their chemical space, and modifies their properties and/or activities. Indole prenyltransferases (IPTs) catalyze the direct regiospecific installation of prenyl moieties on indole-derived compounds. IPTs have shown relaxed substrate flexibility enabling them to be used as tools for indole functionalization. However, the mechanism by which certain IPTs target a specific carbon position is not fully understood. Herein, we use structure-guided site-directed mutagenesis, in vitro enzymatic reactions, kinetics and structural-elucidation of analogs to verify the key catalytic residues that control the regiospecificity of all characterized regiospecific C6 IPTs. The presented results also demonstrate that substitution of PriB_His312 to Tyr leads to the synthesis of analogs prenylated at different positions than C6. This work contributes to understanding of how certain IPTs can access a challenging position in indole-derived compounds.

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