Enzymatic Ring Contraction for the Biosynthesis of Sulfur-Containing Cyclopentachromone

Cyclopentachromone, distinguished by its 6/6/5 heterotricyclic ring structure, is a key building block in many bioactive natural products, yet its enzymatic origin remains unclear. We identified a new class of cyclopentachromone-containing compounds, termed isochromosulfines, characterized by unique C–S bonds. A distinct FAD-dependent monooxygenase, IscL, was identified to catalyze the formation of the 6/6/5 cyclopentadiene intermediate, 2S-remisporine A, from a 6/6/6 xanthone precursor via benzene ring contraction. The high reactivity of 2S-remisporine A further promotes a spontaneous thiol-Michael addition reaction with thiol-containing compounds, forming the C–S bond in isochromosulfines. Additionally, we demonstrate that IscL homologues mediate a bifurcated pathway of benzene ring modification in the xanthone intermediate, leading to either ring contraction or cleavage, which is determined by a critical residue at position 230 to be phenylalanine or tyrosine. Our findings highlight the pivotal role of IscL in forming the 6/6/5 cyclopentachromone scaffold and offer deep insights into its catalytic mechanism. Our work lays the foundation for genome mining of cyclopentachromone-containing compounds and shows the potential application of IscL in biocatalysis.