The structural and functional investigation into an unusual nitrile synthase

Area of Science:

• Biochemistry and Enzymology
• Structural Biology
• Neuroscience

Background:

• The neurotoxin aetokthonotoxin (AETX) possesses a unique pentabrominated biindole nitrile structure.
• AetD is a newly identified nitrile synthase responsible for AETX biosynthesis, exhibiting novel catalytic mechanisms and low sequence homology to known enzymes.

Purpose of the Study:

• To determine the crystal structure of AetD in complex with its substrate.
• To elucidate the substrate-binding pattern and the catalytic mechanism of AetD.
• To investigate the role of substrate bromination in the AetD-catalyzed reaction.

Main Methods:

• X-ray crystallography was employed to resolve the crystal structure of AetD.
• Structures were determined for AetD in complex with various substrates: 5,7-di-Br-L-Trp, 5-Br-L-Trp, and L-Trp.

Main Results:

• AetD adopts a heme oxygenase-like fold, featuring a hydrophobic cavity for indole accommodation.
• A diiron cluster within the active site binds to the carboxyl oxygen and amino nitrogen of the substrate.
• The AetD-catalyzed reaction proceeds independently of substrate bromination.

Conclusions:

• The study reveals the substrate-binding mode and validates the diiron cluster active center of AetD.
• These findings provide a foundational basis for further mechanistic studies of this unique nitrile synthase class.