Cytochrome P450 as dimerization catalyst in diketopiperazine alkaloid biosynthesis

Area of Science:

• Natural Product Biosynthesis
• Microbial Secondary Metabolism
• Enzymology

Background:

• Dimeric natural products often possess significant biological activities.
• Understanding the biosynthesis and dimerization mechanisms of these compounds is crucial for drug discovery.
• (−)-ditryptophenaline, an alkaloid from Aspergillus flavus, shows potential as an analgesic and anti-inflammatory agent by inhibiting the substance P receptor.

Purpose of the Study:

• To elucidate the gene cluster and biosynthetic pathway of the dimeric diketopiperazine alkaloid (−)-ditryptophenaline.
• To identify the enzyme responsible for the formation and dimerization of (−)-ditryptophenaline.
• To investigate the substrate specificity of the identified enzyme.

Main Methods:

• Targeted gene knockout in Aspergillus flavus.
• Heterologous gene expression in yeast.
• Biochemical characterization of the key enzyme.

Main Results:

• The complete gene cluster and biosynthetic pathway for (−)-ditryptophenaline were determined.
• A single cytochrome P450 enzyme, DtpC, was identified as responsible for both pyrroloindole ring formation and monomer dimerization.
• DtpC demonstrated relaxed substrate specificity, enabling the synthesis of novel dimeric compounds from brevianamide F.

Conclusions:

• The study reveals the genetic basis for (−)-ditryptophenaline biosynthesis.
• DtpC is a unique enzyme catalyzing both core structure formation and dimerization.
• A radical-mediated mechanism is proposed for the dimerization process, offering insights into natural product synthesis.