# In vivo and in vitro investigations provide insights into maleidride biosynthesis in fungi

## TL;DR

This paper explores how fungi produce maleidrides, a type of natural compound, and highlights the role of isochorismatase-like enzymes in their biosynthesis.

## Contribution

The study reveals a supportive role of isochorismatase-like enzymes in α-ketoglutarate dependent dioxygenase catalysis during maleidride biosynthesis.

## Key findings

- Gene deletions and enzyme assays confirm the role of isochorismatase-like enzymes in zopfiellin and scytalidin biosynthesis.
- Feeding studies and structural analysis confirm the formation of castaneiolide from scytalidin or its analogue.
- A putative gene cluster for castaneiolide biosynthesis was identified from Macrophoma castaneicola's genome.

## Abstract

Maleidrides are a family of polyketide-derived natural products isolated from filamentous fungi, that can exhibit significant bioactivities. These compounds are classified according to the size of their central carbocyclic ring, to which one or more maleic anhydride moieties are attached. The studies described herein provide important insights into maleidride biosynthesis, in particular the pathways to the nonadrides scytalidin and castaneiolide, and the octadride zopfiellin. We propose a supportive role for isochorismatase-like enzymes, which are commonly encoded within maleidride biosynthetic gene clusters, in facilitating α-ketoglutarate dependent dioxygenase-mediated catalysis. This is evidenced by gene deletions as well as enzyme assays, for two maleidride biosynthetic pathways: that of zopfiellin, from Diffractella curvata; and of scytalidin, from Scytalidium album. These experiments collectively underscore the significance of the isochorismatase-like enzymes in the catalytic process of α-ketoglutarate dependent dioxygenases. Feeding studies with either scytalidin or an unsaturated analogue to D. curvata ΔzopPKS both gave the 5,6-diol, castaneiolide and the structure was confirmed by NMR and X-ray crystallography. Furthermore, a putative biosynthetic gene cluster for castaneiolide biosynthesis was identified from a de novo genome assembly of the native producer, Macrophoma castaneicola.

Isochorismatase-like enzymes play a supportive role in α-ketoglutarate dependent dioxygenase mediated catalysis to increase maleidride diversity.