Crystal structure and functional analysis of large-terpene synthases belonging to a newly found subclass

Fujihashi, M.; Sato, Tsutomu; Tanaka, Yuma; Yamamoto, Daisuke; Nishi, Tohru; Ueda, Daijiro; Murakami, Mizuki; Yasuno, Yoko; Sekihara, Ai; Fuku, Kazuma; Shinada, Tetsuro; Miki, Kunio

doi:10.1039/c8sc00289d

Cited by 31 publications

(52 citation statements)

References 16 publications

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“…2C). The active site arrangement of DabA is also distinct from the recently described class IB terpene synthases that are responsible for the biosynthesis of large terpene products (C 25 /C 30 /C 35 ) (25). Class IB terpene synthases lack the canonical NSE and DDxxD motifs.…”

Section: Resultsmentioning

confidence: 86%

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

Chekan

McKinnie

Noel

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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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.

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Section: Resultsmentioning

confidence: 86%

“…Class IB terpene synthases lack the canonical NSE and DDxxD motifs. Instead, they have new aspartate rich motifs that occupy the same 3D location within the active site, even though they do not align in primary sequence comparisons (25). This contrasts with DabA which retains the NSE motif but completely lacks the DDxxD motif or any substitute.…”

Section: Resultsmentioning

confidence: 96%

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

Chekan

McKinnie

Noel

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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show abstract

“…The active site arrangement of DabA is also distinct from the recently described class IB terpene synthases that are responsible for the biosynthesis of large terpene products (C25/C30/C35) (24). Class IB terpene synthases lack the canonical NSE and DDxxD motifs, but have replaced them with new aspartate rich motifs that occupy the same location within the active site (24). This contrasts with DabA which retains the NSE motif, but lacks the DDxxD motif or any substitute.…”

Section: Active Sitementioning

confidence: 82%

“…Examination of the crystal structure shows that the canonical DDxxD motif has not been replaced by another series of magnesium binding residues; no magnesium is present in this region of the active site ( Figure 2C). The active site arrangement of DabA is also distinct from the recently described class IB terpene synthases that are responsible for the biosynthesis of large terpene products (C25/C30/C35) (24). Class IB terpene synthases lack the canonical NSE and DDxxD motifs, but have replaced them with new aspartate rich motifs that occupy the same location within the active site (24).…”

Section: Active Sitementioning

confidence: 86%

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

Chekan

McKinnie

Noel

et al. 2020

Preprint

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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 Nprenylation 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 Mg 2+ -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 Nprenylation 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.

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“…Interestingly, the overall structure of BalTS is similar to that of group I terpene synthases, but it consists of two novel aspartate-rich motifs, DYLDNLxD and DY(F,L,W)IDxxED, which forms the cyclic skeleton. 102 More recently, Stepanova et al also determined the crystal structure of BalTS in a complex with a substrate surrogate and elucidated the catalytic mechanisms in unusual large terpene synthesis and in drug design applications. On the basis of these findings, the authors designated 35-carbon terpene as a new family named sesquarterpenes.…”

Section: New Subclass: Sesquarterpene Cyclasementioning

confidence: 99%

Structure, catalysis, and inhibition mechanism of prenyltransferase

Chang¹,

Cheng²,

Wang

2020

IUBMB Life

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Isoprenoids, also known as terpenes or terpenoids, represent a large family of natural products composed of five-carbon isopentenyl diphosphate or its isomer dimethylallyl diphosphate as the building blocks. Isoprenoids are structurally and functionally diverse and include dolichols, steroid hormones, carotenoids, retinoids, aromatic metabolites, the isoprenoid side-chain of ubiquinone, and isoprenoid attached signaling proteins. Productions of isoprenoids are catalyzed by a group of enzymes known as prenyltransferases, such as farnesyltransferases, geranylgeranyltransferases, terpenoid cyclase, squalene synthase, aromatic prenyltransferase, and cis-and trans-prenyltransferases. Because these enzymes are key in cellular processes and metabolic pathways, they are expected to be potential targets in new drug discovery. In this review, six distinct subsets of characterized prenyltransferases are structurally and mechanistically classified, including (1) head-to-tail prenyl synthase, (2) head-to-head prenyl synthase, (3) head-tomiddle prenyl synthase, (4) terpenoid cyclase, (5) aromatic prenyltransferase, and (6) protein prenylation. Inhibitors of those enzymes for potential therapies against several diseases are discussed. Lastly, recent results on the structures of integral membrane enzyme, undecaprenyl pyrophosphate phosphatase, are also discussed.

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Crystal structure and functional analysis of large-terpene synthases belonging to a newly found subclass

Abstract: The first crystal structure of a large-terpene synthase elucidated two novel Asp rich motifs and a new subclass of terpene synthases.

Cited by 31 publications

References 16 publications

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

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

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

Structure, catalysis, and inhibition mechanism of prenyltransferase

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