California Sea Grant College Program

The marine sediment-derived Streptomyces spp. CNQ-525 and CNQ-766 were recently characterized by the Fenical laboratory as a new group of marine sediment-derived actinomycetes, tentatively named the MAR4s. These bacteria are prolific producers of hybrid isoprenoids, including the meroterpenoid (polyketide-terpene) antibiotics that are rarely encountered from bacteria. Structural inspection of the meroterpenoid antibiotics belonging to the napyradiomycin family of chlorinated dihydroquinones suggests that the biosynthetic cyclization of their terpenoid subunits is initiated via a chloronium ion, which exists as hypochlorous acid. The vanadium-dependent haloperoxidases that are known to catalyze such reactions are distributed in fungi and marine algae and have yet to be characterized from bacteria.

The MAR4 strain Streptomyces sp. CNQ-525 was used as a source for identifying novel halogenating enzymes. The cloning and sequence analysis of the 43-kb napyradiomycin biosynthetic cluster yielded an unprecedented arrangement of biosynthetic genes including a FADH₂-dependent halogenase and three putative vanadium-dependent chloroperoxidases. Heterologous expression of the CNQ-525-based nap biosynthetic cluster in a surrogate host produced at least seven napyradiomycins, including the new analog 2-deschloro- 2-hydroxy-A80915C. These data revealed the molecular basis behind the biosynthesis of these novel meroterpenoid natural products, and also resulted in the first identification of vanadium-dependent haloperoxidases from a prokaryote. Preliminary biochemical data suggests both NapH1 and NapH3 from the napyradiomycin biosynthetic cluster are vanadium-dependent chloroperoxidases and inactivation of the napH1 gene resulted in the first in vivo verification of a vanadium-dependent haloperoxidase.

The phthalazinone meroterpenoid azamerone was isolated by the Fenical- Jensen lab from the marine sediment-derived bacterium Streptomyces sp. CNQ- 766. Given the structural novelty of azamerone and its relation to the chlorinated meroterpenoids, a variety of feeding experiments with ¹³C and ¹⁵N-labeled molecules were used to explore the formation of its phthalazinone core. These experiments confirmed that azamerone’s phthalazinone core is derived from a naphthoquinone precursor such as in the chlorinated dihydroquinones and progresses through a diazo intermediate.