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

Abstract

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 (<i>cyb</i>) is identified by targeted bioinformatic screening and validated by heterologous expression in <i>E. coli</i>. Functional evaluation of the recombinant key enzymes provides in-depth mechanistic insights into a streamlined <i>C</i>,<i>C</i>-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 <i>in vitro</i> 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.