Heterodimeric Non-heme Iron Enzymes in Fungal Meroterpenoid Biosynthesis.
Xinyang LiTakayoshi AwakawaTakahiro MoriMeiqi LingDan HuBin WuLeonard KaysserPublished in: Journal of the American Chemical Society (2021)
Talaromyolides (1-6) are a group of unusual 6/6/6/6/6/6 hexacyclic meroterpenoids with (3R)-6-hydroxymellein and 4,5-seco-drimane substructures, isolated from the marine fungus Talaromyces purpureogenus. We have identified the biosynthetic gene cluster tlxA-J by heterologous expression in Aspergillus, in vitro enzyme assays, and CRISPR-Cas9-based gene inactivation. Remarkably, the heterodimer of non-heme iron (NHI) enzymes, TlxJ-TlxI, catalyzes three steps of oxidation including a key reaction, hydroxylation at C-5 and C-9 of 12, the intermediate with 3-ketohydroxydrimane scaffold, to facilitate a retro-aldol reaction, leading to the construction of the 4,5-secodrimane skeleton and characteristic ketal scaffold of 1-6. The products of TlxJ-TlxI, 1 and 4, were further hydroxylated at C-4'β by another NHI heterodimer, TlxA-TlxC, and acetylated by TlxB to yield the final products, 3 and 6. The X-ray structural analysis coupled with site-directed mutagenesis provided insights into the heterodimer TlxJ-TlxI formation and its catalysis. This is the first report to show that two NHI proteins form a heterodimer for catalysis and utilizes a novel methodology to create functional oxygenase structures in secondary metabolite biosynthesis.
Keyphrases
- crispr cas
- cell wall
- genome editing
- copy number
- high resolution
- genome wide
- poor prognosis
- visible light
- genome wide identification
- tissue engineering
- electron transfer
- hydrogen peroxide
- magnetic resonance imaging
- dna methylation
- mass spectrometry
- high throughput
- gene expression
- genome wide analysis
- nitric oxide
- binding protein
- single cell
- magnetic resonance
- saccharomyces cerevisiae
- electron microscopy