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Pneumocandins produced by the fungusGlarea lozoyensis are acylated cyclic hexapeptides of the echinocandin family. Pneumocandin B0 is the starting molecule for the first semisynthetic echinocandin antifungal drug, caspofungin acetate. In the wild-type strain, pneumocandin B0 is a minor fermentation product, and its industrial production was achieved by a combination of extensive mutation and medium optimization. The pneumocandin biosynthetic gene cluster was previously elucidated by a whole-genome sequencing approach. Knowledge of the biosynthetic cluster suggested an alternative way to produce exclusively pneumocandin B0 . Disruption ofGLOXY4 , encoding a nonheme, α-ketoglutarate-dependent oxygenase, confirmed its involvement inl -leucine cyclization to form 4S -methyl-l -proline. The absence of 4S -methyl-l -proline abolishes pneumocandin A0 production, and 3S -hydroxyl-l -proline occupies the hexapeptide core's position 6, resulting in exclusive production of pneumocandin B0 . Retrospective analysis of theGLOXY4 gene in a previously isolated pneumocandin B0 -exclusive mutant (ATCC 74030) indicated that chemical mutagenesis disrupted theGLOXY4 gene function by introducing two amino acid mutations in GLOXY4. This one-step genetic manipulation can rationally engineer a high-yield production strain.

Engineering of Glarea lozoyensis for Exclusive Production of the Pneumocandin B 0 Precursor of the Antifungal Drug Caspofungin Acetate

Engineering of Glarea lozoyensis for Exclusive Production of the Pneumocandin B ₀ Precursor of the Antifungal Drug Caspofungin Acetate