Unveiling the Bacterial Sesquiterpenome of Streptomyces sp. CBMAI 2042 Discloses Cyclases with Versatile Performances

Terpenes are the most abundant class of natural product that exist in nature. They possess a myriad of industrial applications including pharmaceutical, perfumery and flavors, bulk chemicals, and fuel. Intriguingly, until today, the vast majority of characterized terpenoids have been isolated from plants and fungi, and only in recent years bacteria were found to generate a representative reservoir of terpenoids molecules. Mining Streptomyces sp. CBMAI 2042 genome data has revealed the presence of five terpene cyclase genes. Chemical analysis of mycelium extract of this bacteria strain has unveiled at least 28 volatile terpenes molecules, where three encoding sesquiterpene cyclase (STC) genes are apparently responsible for their biosynthesis. The cyclic products obtained by incubation of these three purified recombinant STCs with farnesyl diphosphate (FPP) were analyzed by gas chromatography-mass spectrometry (GC-MS) and identified using the Van den Dool and Kratz equation.

Computational Studies on the Sterol-like Cyclization of the Monodomain Class II Terpene Cyclase

The proposed mechanism for the sterol-like cyclization of a monodomain class II terpene cyclase was studied computationally by using density functional theory (DFT). The complete pathway for the conversion of...

Algal neurotoxin biosynthesis repurposes the terpene cyclase structural fold into anN-prenyltransferase

Prenylation is a common biological reaction in all domains of life wherein prenyl diphosphate donors transfer prenyl groups onto small molecules as well as large proteins. The enzymes that catalyze these reactions are structurally distinct from ubiquitous terpene cyclases that, instead, assemble terpenes via intramolecular rearrangements of a single substrate. Herein, we report the structure and molecular details of a new family of prenyltransferases from marine algae that repurposes the terpene cyclase structural fold for theN-prenylation of glutamic acid during the biosynthesis of the potent neurochemicals domoic acid and kainic acid. We solved the X-ray crystal structure of the prenyltransferase found in domoic acid biosynthesis, DabA, and show distinct active site binding modifications that remodel the canonical magnesium (Mg2+)-binding motif found in terpene cyclases. We then applied our structural knowledge of DabA and a homologous enzyme from the kainic acid biosynthetic pathway, KabA, to reengineer their isoprene donor specificities (geranyl diphosphate [GPP] versus dimethylallyl diphosphate [DMAPP]) with a single amino acid change. While diatom DabA and seaweed KabA enzymes share a common evolutionary lineage, they are distinct from all other terpene cyclases, suggesting a very distant ancestor to the larger terpene synthase family.

Algal Neurotoxin Biosynthesis Repurposes the Terpene Cyclase Structural Fold Into an N-prenyltransferase

Prenylation is a common biological reaction in all domains of life whereupon prenyl diphosphate donors transfer prenyl groups onto small molecules as well as large proteins. The enzymes that catalyze these biotransformations are structurally distinct from ubiquitous terpene cyclases that instead assemble terpene molecules via intramolecular rearrangements. Herein we report the structure and molecular details of a new family of prenyltransferases from marine algae that repurposes the terpene cyclase structural fold for the N-prenylation of glutamic acid during the biosynthesis of the potent neurochemicals domoic acid and kainic acid. We solved the X-ray crystal structure of the prenyltransferase found in domoic acid biosynthesis, DabA, and show distinct active site binding modifications that remodel the canonical Mg2+-binding motif. We then applied our structural knowledge of DabA and a homologous enzyme from the kainic acid biosynthetic pathway, KabA, to alter their isoprene donor specificities (geranyl versus dimethylallyl diphosphate) by a single amino acid switch. While the diatom DabA and seaweed KabA enzymes share a common evolutionary lineage, they are distinct from all other terpene cyclases, suggesting a very distant ancestor.SignificanceDomoic acid is a neurotoxin produced by marine algae that readily bioaccumulates in shellfish and significantly impacts both human and animal life. The first committed step of the biosynthesis of domoic acid is the N-prenylation of L-glutamic acid by the enzyme DabA. By solving the crystal structure of DabA, we demonstrate that this enzyme has repurposed the common terpene cyclase fold to catalyze an extremely unusual reaction, N-prenylation of an unactivated primary amine. Application of these structural insights enabled rational engineering of two N-prenyltransferase enzymes to accept alternative prenyl donors. Ultimately, these results not only expand the scope of reactions catalyzed by a terpene cyclase family member, but will help inform future domoic acid environmental monitoring efforts.