Biosynthesis of Neocarazostatin A Reveals the Sequential Carbazole Prenylation and Hydroxylation in the Tailoring Steps

Abstract

Highlights
• The biosynthetic gene cluster of neocarazostatin A was identified
• A new type of carbazole prenyltransferases, NzsG, was characterized
• The P450 enzyme NzsA catalyzing the last step of the biosynthesis was identified
• The biotransformation in the late stage of the biosynthesis was reconstituted

Summary
Neocarazostatin A (NZS) is a bacterial alkaloid with promising bioactivities against free radicals, featuring a tricyclic carbazole nucleus with a prenyl moiety at C-6 of the carbazole ring. Here, we report the discovery and characterization of the biosynthetic pathway of NZS through genome mining and gene inactivation. The in vitro assays characterized two enzymes: NzsA is a P450 hydroxylase and NzsG is a new phytoene-synthase-like prenyltransferase (PTase). This is the first reported native PTase that specifically acts on the carbazole nucleus. Finally, our in vitro reconstituted experiment demonstrated a coupled reaction catalyzed by NzsG and NzsA tailoring the NZS biosynthesis.