Discovery, Structure, and Mechanism of a Class II Sesquiterpene Cyclase

Abstract

Terpene cyclases (TCs), extraordinary enzymes that create the structural diversity seen in terpene natural products, are traditionally divided into two classes, class I and class II. Although the structural and mechanistic features of class I TCs are well-known, the corresponding details in class II counterparts have not been fully characterized. Here, we report the genome mining discovery and structural characterization of two class II sesquiterpene cyclases (STCs) from Streptomyces. These drimenyl diphosphate synthases (DMSs) are the first STCs shown to possess β,γ-didomain architecture. High-resolution X-ray crystal structures of DMS from Streptomyces showdoensis (SsDMS) in complex with both a farnesyl diphosphate and Mg2+ unveiled an induced-fit mechanism, with an unprecedented Mg2+ binding mode, finally solving one of the lingering questions in class II TC enzymology. This study supports continued genome mining for novel bacterial TCs and provides new mechanistic insights into canonical class II TCs that will lead to advances in TC engineering and synthetic biology.

Isolation, (bio)synthetic studies and evaluation of antimicrobial properties of drimenol-type sesquiterpenes of Termitomyces fungi

“…The biosynthesis of drimenol ( 1 ) has also gained much attention as drimenol can be formed via both class I and class II catalytic mechanisms. While drimenol synthases identified from plants were characterized as class I terpene cyclases (TCs), bacterial drimenol synthases were identified as class II sesquiterpene cyclases 50 – 52 . In contrast, dedicated bifunctional haloacid dehalogenase-like (HAD-like) terpene cyclases were reported from Aspergillus species 53 , 54 and most recently from Antrodia cinnamomea 55 .…”

Terpenoids from Penicillium chrysogenum MT-40, an endophytic fungus isolated from Huperzia serrata

“…Notably, although a handful of TPCs such as TnlT2 and Bra4 in the biosynthesis of the antibiotic tiancilactone A and the immunosuppressor brasilicardin A can employ epoxy GGPP to generate diterpene skeleton with 3/3/3 fusing ring system, [21][22][23] all the known TPCs involved in the biosynthesis of drimane-type sesquiterpenoids utilize FPP rather than epoxy FPP as substrate to perform the cascade cyclization reaction. [24][25][26] Supposedly, there might be a functionally noncanonical TPC in P. chrysogenum MT-40 playing crucial role in the biosynthesis of 4 from epoxy FPP, and the further characterization of this noncanonical TPC will undoubtedly deepen the understanding of the biosynthesis of driane-type sesquiterpenoids.…”

Terpene Synthases in the Biosynthesis of Drimane‐Type Sesquiterpenes across Diverse Organisms

“…The conserved DxDD motif or DxDT motif, characteristic of class II TPSs, can be found in SsDMS [17] and AsDMS [14] from bacteria, as well as in AstC, [11] DtrB, [12] AncC, [13] AncA, [19] and DS3 [15] from fungi (Figure 5A). Mutagenesis studies were conducted targeting the second aspartic acid residue in SsDMS (DxD 303 D to DxA 303 D), AsDMS (DxD 333 T to DxN 333 T), and AncC (DxD 288 T to DxA 288 T), resulting in diminished production of drimane products.…”

“…SsDMS from S. showdoensis is the first structurally elucidated drimenyl pyrophosphate synthase, featuring a βγ ‐didomain architecture with the catalytic DxDD motif located at the β ‐ γ interface [17] . X‐ray crystal structures of SsDMS with FPP and Mg 2+ unveil an induced‐fit mechanism and unique Mg 2+ binding.…”

“…Two dephosphorylases, AstI and AstK, belonging to the HAD-like hydrolase superfamily, catalyze the successive dephosphorylation of drimenyl pyrophosphate (13) to produce drim-8-ene-11-ol (9) via drimenyl monophosphate (14). Furthermore, the non-ribosomal peptide synthetase AstA and acetyltransferase AstG play roles in esterifying aryl acids (benzoic acid or 4-hydroxybenzoic acid) and acetic acid, respectively, to generate DTS lactones (16)(17)(18)(19).…”