(Bio-)Synthesis of the Aquatic Phytotoxin Cyanobacterin – A Paradigm for Furanolide Core Structure Assembly

The gamma-butyrolactone structural motif is commonly found in many natural signaling molecules and other specialized metabolites. A prominent example is the potent aquatic phytotoxin cyanobacterin bearing a highly functionalized gamma-butyrolactone core structure. The enzymatic machinery assembling cyanobacterin and the many structurally related natural products – herein termed furanolides – has remained elusive over decades. Here we discover and characterize the underlying biosynthetic process of furanolide core structure assembly. The cyanobacterin biosynthetic gene cluster (cyb) is identified by targeted bioinformatic screening and validated by heterologous expression in E. coli. Functional evaluation of the recombinant key enzymes provides in-depth mechanistic insights into a streamlined C,C-bond-forming cascade that involves installation of compatible reactivity at seemingly unreactive C-alpha-positions of the amino acid precursors and facilitates development of a one-pot biocatalytic in vitro synthesis. Our work extends the biosynthetic and biocatalytic toolbox for gamma-butyrolactone formation. It thereby provides a general paradigm for the biosynthesis of furanolides and thus sets the stage for their targeted discovery, biosynthetic engineering and enzymatic synthesis.