An ATP-dependent ligase catalyzes the fourth ring cyclization in tetracycline biosynthesis

Abstract

Cyclization steps in biosynthesis of aromatic polyketides are typically directed by specific enzymes known as cyclases. The fourth ring cyclization step that forms the fully aromatic pretetramid during tetracycline biosynthesis has not been resolved. Herein we report in vitro and in vivo studies on the fourth ring cyclization step in biosynthesis of SF2575 and related tetracyclic natural products. We demonstrate that an enzyme belonging to the ATP-dependent acyl-CoA ligase family catalyzes the C1–C18 Claisen condensation step that forms the A ring and yields the linear, tetracyclic aromatic compound that is the precursor to tetracyclic natural products. The enzyme, SsfL2, is well-conserved in all tetracycline biosynthetic gene clusters discovered to date. It is proposed that SsfL2 directly adenylates the tricyclic carboxylic acid to facilitate the final carbon-carbon bond formation, and coenzyme A is not required.