Terpenoid synthase structures: a so far incomplete view of complex catalysis

Terpenoid synthases are ubiquitous enzymes that catalyze the formation of structurally and stereochemically diverse isoprenoid natural products. Many isoprenoid coupling enzymes and terpenoid cyclases from bacteria, fungi, protists, plants, and animals share the class I terpenoid synthase fold. Despite generally low amino acid sequence identity among these examples, class I terpenoid synthases contain conserved metal-binding motifs that coordinate to a trinuclear metal cluster. This cluster not only serves to bind and orient the flexible isoprenoid substrate in the precatalytic Michaelis complex, but it also triggers the departure of the diphosphate leaving group to generate a carbocation that initiates catalysis. Additional conserved hydrogen bond donors assist the metal cluster in this function. Crystal structure analysis reveals that the constellation of three metal ions required for terpenoid synthase catalysis is generally identical among all class I terpenoid synthases of known structure.

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Investigation of terpene diversification across multiple sequenced plant genomes

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1419547112 ◽

2014 ◽

Vol 112 (1) ◽

pp. E81-E88 ◽

Cited By ~ 137

Author(s):

Alexander M. Boutanaev ◽

Tessa Moses ◽

Jiachen Zi ◽

David R. Nelson ◽

Sam T. Mugford ◽

...

Keyword(s):

Cytochromes P450 ◽

Gene Clusters ◽

Metabolic Diversity ◽

Plant Genomes ◽

Gene Pairs ◽

Genetic Components ◽

Dynamic Genome ◽

Scaffold Diversity ◽

Terpenoid Synthase ◽

Key Events

Plants produce an array of specialized metabolites, including chemicals that are important as medicines, flavors, fragrances, pigments and insecticides. The vast majority of this metabolic diversity is untapped. Here we take a systematic approach toward dissecting genetic components of plant specialized metabolism. Focusing on the terpenes, the largest class of plant natural products, we investigate the basis of terpene diversity through analysis of multiple sequenced plant genomes. The primary drivers of terpene diversification are terpenoid synthase (TS) “signature” enzymes (which generate scaffold diversity), and cytochromes P450 (CYPs), which modify and further diversify these scaffolds, so paving the way for further downstream modifications. Our systematic search of sequenced plant genomes for all TS and CYP genes reveals that distinct TS/CYP gene pairs are found together far more commonly than would be expected by chance, and that certain TS/CYP pairings predominate, providing signals for key events that are likely to have shaped terpene diversity. We recover TS/CYP gene pairs for previously characterized terpene metabolic gene clusters and demonstrate new functional pairing of TSs and CYPs within previously uncharacterized clusters. Unexpectedly, we find evidence for different mechanisms of pathway assembly in eudicots and monocots; in the former, microsyntenic blocks of TS/CYP gene pairs duplicate and provide templates for the evolution of new pathways, whereas in the latter, new pathways arise by mixing and matching of individual TS and CYP genes through dynamic genome rearrangements. This is, to our knowledge, the first documented observation of the unique pattern of TS and CYP assembly in eudicots and monocots.

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Genomic Organization of Plant Terpene Synthases and Molecular Evolutionary Implications

Genetics ◽

10.1093/genetics/158.2.811 ◽

2001 ◽

Vol 158 (2) ◽

pp. 811-832

Author(s):

Susan C Trapp ◽

Rodney B Croteau

Keyword(s):

Natural Products ◽

Secondary Metabolism ◽

Intron Loss ◽

Terpene Synthase ◽

Primary Metabolism ◽

Taxus Brevifolia ◽

Terpene Synthases ◽

Sequence Element ◽

Geranyl Diphosphate ◽

Terpenoid Synthase

Abstract Terpenoids are the largest, most diverse class of plant natural products and they play numerous functional roles in primary metabolism and in ecological interactions. The first committed step in the formation of the various terpenoid classes is the transformation of the prenyl diphosphate precursors, geranyl diphosphate, farnesyl diphosphate, and geranylgeranyl diphosphate, to the parent structures of each type catalyzed by the respective monoterpene (C10), sesquiterpene (C15), and diterpene synthases (C20). Over 30 cDNAs encoding plant terpenoid synthases involved in primary and secondary metabolism have been cloned and characterized. Here we describe the isolation and analysis of six genomic clones encoding terpene synthases of conifers, [(-)-pinene (C10), (-)-limonene (C10), (E)-α-bisabolene (C15), δ-selinene (C15), and abietadiene synthase (C20) from Abies grandis and taxadiene synthase (C20) from Taxus brevifolia], all of which are involved in natural products biosynthesis. Genome organization (intron number, size, placement and phase, and exon size) of these gymnosperm terpene synthases was compared to eight previously characterized angiosperm terpene synthase genes and to six putative terpene synthase genomic sequences from Arabidopsis thaliana. Three distinct classes of terpene synthase genes were discerned, from which assumed patterns of sequential intron loss and the loss of an unusual internal sequence element suggest that the ancestral terpenoid synthase gene resembled a contemporary conifer diterpene synthase gene in containing at least 12 introns and 13 exons of conserved size. A model presented for the evolutionary history of plant terpene synthases suggests that this superfamily of genes responsible for natural products biosynthesis derived from terpene synthase genes involved in primary metabolism by duplication and divergence in structural and functional specialization. This novel molecular evolutionary approach focused on genes of secondary metabolism may have broad implications for the origins of natural products and for plant phylogenetics in general.

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A Comparison of Two Monoterpenoid Synthases Reveals Molecular Mechanisms Associated With the Difference of Bioactive Monoterpenoids Between Amomum villosum and Amomum longiligulare

Frontiers in Plant Science ◽

10.3389/fpls.2021.695551 ◽

2021 ◽

Vol 12 ◽

Author(s):

Haiying Zhao ◽

Meng Li ◽

Yuanyuan Zhao ◽

Xiaojing Lin ◽

Huilin Liang ◽

...

Keyword(s):

Molecular Mechanisms ◽

Binding Pocket ◽

Major Product ◽

Bornyl Acetate ◽

Directed Mutation ◽

Key Enzyme ◽

Volatile Terpenoids ◽

The Difference ◽

Terpenoid Synthase ◽

Amomum Villosum

The fruits of Amomum villosum and Amomum longiligulare are both used medicinally as Fructus Amomi the famous traditional Chinese medicine, however, the medicinal quality of A. villosum is better than that of A. longiligulare. Volatile terpenoids in the seeds, especially bornyl acetate and borneol, are the medicinal components of Fructus Amomi. The volatile terpenoids and transcriptome of developing seeds of A. villosum and A. longiligulare were compared in this study. The result revealed that the bornyl acetate and borneol contents were higher in A. villosum than in A. longiligulare. Additionally, six terpenoid synthase genes (AlTPS1–AlTPS6) were screened from the transcriptome of A. longiligulare, and AlTPS2 and AlTPS3 were found to share 98 and 83% identity with AvTPS2 and AvBPPS (bornyl diphosphate synthase) from A. villosum, respectively. BPPS is the key enzyme for the biosynthesis of borneol and bornyl acetate. Biochemical assays improved that AlTPS2 had an identical function to AvTPS2 as linalool synthase; however, AlTPS3 produced camphene as the major product and bornyl diphosphate (BPP) as the secondary product, whereas AvBPPS produced BPP as its major product. There was only one different amino acid between AlTPS3 (A496) and AvBPPS (G495) in their conserved motifs, and the site-directed mutation of A496G in DTE motif of AlTPS3 changed the major product from camphene to BPP. Molecular docking suggests that A496G mutation narrows the camphene-binding pocket and decreases the BPP-binding energy, thus increases the product BPP selectivity of enzyme. In addition, the expression level of AvBPPS was significantly higher than that of AlTPS3 in seeds, which was consistent with the related-metabolites contents. This study provides insight into the TPS-related molecular bases for the biosynthesis and accumulation differences of the bioactive terpenoids between A. villosum and A. longiligulare. BPPS, the key gene involved in the biosynthesis of the active compound, was identified as a target gene that could be applied for the quality-related identification and breeding of Fructus Amomi.

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Identification and Functional Characterization of the Gene Cluster Responsible for Fusaproliferin Biosynthesis in Fusarium proliferatum

Toxins ◽

10.3390/toxins13070468 ◽

2021 ◽

Vol 13 (7) ◽

pp. 468

Author(s):

Asja Ćeranić ◽

Thomas Svoboda ◽

Franz Berthiller ◽

Michael Sulyok ◽

Jonathan Matthew Samson ◽

...

Keyword(s):

Fusarium Species ◽

Functional Characterization ◽

Fusarium Proliferatum ◽

Cytochrome P450s ◽

Gene Encoding ◽

Knock Out ◽

Genes Encoding ◽

Terpenoid Synthase ◽

Taxonomic Groups

The emerging mycotoxin fusaproliferin is produced by Fusarium proliferatum and other related Fusarium species. Several fungi from other taxonomic groups were also reported to produce fusaproliferin or the deacetylated derivative, known as siccanol or terpestacin. Here, we describe the identification and functional characterization of the Fusarium proliferatum genes encoding the fusaproliferin biosynthetic enzymes: a terpenoid synthase, two cytochrome P450s, a FAD-oxidase and an acetyltransferase. With the exception of one gene encoding a CYP450 (FUP2, FPRN_05484), knock-out mutants of the candidate genes could be generated, and the production of fusaproliferin and intermediates was tested by LC-MS/MS. Inactivation of the FUP1 (FPRN_05485) terpenoid synthase gene led to complete loss of fusaproliferin production. Disruption of a putative FAD-oxidase (FUP4, FPRN_05486) did not only affect oxidation of preterpestacin III to terpestacin, but also of new side products (11-oxo-preterpstacin and terpestacin aldehyde). In the knock-out strains lacking the predicted acetyltransferase (FUP5, FPRN_05487) fusaproliferin was no longer formed, but terpestacin was found at elevated levels. A model for the biosynthesis of fusaproliferin and of novel derivatives found in mutants is presented.

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Stout camphor tree genome fills gaps in understanding of flowering plant genome and gene family evolution

10.1101/371112 ◽

2018 ◽

Cited By ~ 1

Author(s):

Shu-Miaw Chaw ◽

Yu-Ching Liu ◽

Han-Yu Wang ◽

Yu-Wei Wu ◽

Chan-Yi Ivy Lin ◽

...

Keyword(s):

Gene Family Evolution ◽

Plant Genome ◽

Flowering Plant ◽

Small Scale ◽

Phylogenomic Analysis ◽

Segmental Duplications ◽

Duplication Events ◽

Reference Quality ◽

Terpenoid Synthase ◽

Camphor Tree

AbstractWe present reference-quality genome assembly and annotation for the stout camphor tree (SCT; Cinnamomum kanehirae [Laurales, Lauraceae]), the first sequenced member of the Magnoliidae comprising four orders (Laurales, Magnoliales, Canellales, and Piperales) and over 9,000 species. Phylogenomic analysis of 13 representative seed plant genomes indicates that magnoliid and eudicot lineages share more recent common ancestry relative to monocots. Two whole genome duplication events were inferred within the magnoliid lineage, one before divergence of Laurales and Magnoliales and the other within the Lauraceae. Small scale segmental duplications and tandem duplications also contributed to innovation in the evolutionary history of Cinnamomum. For example, expansion of terpenoid synthase subfamilies within the Laurales spawned the diversity of Cinnamomum monoterpenes and sesquiterpenes.

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