Structure Determination and Biosynthesis of Dapalides A–C, Glycosylated Kahalalide F Analogues from the Marine Cyanobacterium <i>Dapis</i> sp

Kahalalides were originally isolated from the marine mollusk <i>Elysia rufescens</i> and its green algal diet <i>Bryopsis</i> sp., but the true producer was revealed as the obligate bacterial symbiont <i>Candidatus</i> Endobryopsis kahalalidefaciens, residing within <i>Bryopsis</i> sp. The most notable is kahalalide F, a broad-spectrum antitumor depsipeptide that entered the clinic but failed from lack of efficacy. We have isolated three new glycosylated analogues of kahalalide F, termed dapalides A-C (<b>1</b>-<b>3</b>), from a marine cyanobacterium, <i>Dapis</i> sp., collected from Guam. The planar structures were determined by extensive NMR coupled with mass spectrometry. Acid hydrolysis of <b>1</b> using amino acid analysis revealed the absolute configuration of singlet and a mixture of duplicate amino acids. Metagenomic analysis unveiled a biosynthetic gene cluster (BGC) with a nonribosomal peptide synthetase (NRPS) system and downstream glycosylation enzymes, which assisted the configurational assignment through epimerization domain analysis. The discovered BGC, termed <i>dap</i>, was assigned to a high-quality metagenome-assembled genome of the <i>Dapis</i> sp. Dapalide A (<b>1</b>) was subjected to phenotypic bioassays and exhibited weak anticancer cytotoxicity. This discovery expands the chemical diversity of the kahalalide F family, suggests their broad ecological role across diverse organisms, and presents an intriguing case of natural product biosynthesis evolution.