The diterpenoids are classically defined by their composition--four isoprenyl units (20 carbons)--and are generally derived from E,E,E-geranylgeranyl diphosphate (GGPP). Such metabolism seems to be ...ancient and has been extensively diversified, with ∼12,000 diterpenoid natural products known. Particularly notable are the gibberellin phytohormones, whose requisite biosynthesis has provided a genetic reservoir that gave rise to not only a large superfamily of ∼7,000 diterpenoids but also, to some degree, all plant terpenoid natural products. This review focuses on the diterpenoids, particularly the defining biosynthetic characteristics of the major superfamilies defined by the cyclization and/or rearrangement of GGPP catalyzed by diterpene synthases/cyclases, although it also includes some discussion of the important subsequent elaboration in the few cases where sufficient molecular genetic information is available. It additionally addresses the array of biological activity providing the selective pressures that drive the observed gene family expansion and diversification, along with biosynthetic gene clustering.
Tanshinones are the bioactive diterpenoid constituents of the traditional Chinese medicinal herb Danshen (Salvia miltiorrhiza), and are examples of the phenolic abietanes widely found within the ...Lamiaceae plant family. Due to the significant interest in these labdane-related diterpenoid natural products, their biosynthesis has been intensively investigated. In addition to providing the basis for metabolic engineering efforts, this work further yielded pioneering insights into labdane-related diterpenoid biosynthesis in the Lamiaceae more broadly. This includes stereochemical foreshadowing of aromatization, with novel protein domain loss in the relevant diterpene synthase, as well as broader phylogenetic conservation of the relevant enzymes. Beyond such summary of more widespread metabolism, formation of the furan ring that characterizes the tanshinones also has been recently elucidated. Nevertheless, the biocatalysts for the pair of demethylations remain unknown, and the intriguing potential connection of these reactions to the further aromatization observed in the tanshinones are speculated upon here.
The complexity of terpenoid natural products has drawn significant interest, particularly since their common (poly)isoprenyl origins were discovered. Notably, much of this complexity is derived from ...the highly variable cyclized and/or rearranged nature of the observed hydrocarbon skeletal structures. Indeed, at least in some cases it is difficult to immediately recognize their derivation from poly-isoprenyl precursors. Nevertheless, these diverse structures are formed by sequential elongation to acyclic precursors, most often with subsequent cyclization and/or rearrangement. Strikingly, the reactions used to assemble and diversify terpenoid backbones share a common carbocationic driven mechanism, although the means by which the initial carbocation is generated does vary. High-resolution crystal structures have been obtained for at least representative examples from each of the various types of enzymes involved in producing terpenoid hydrocarbon backbones. However, while this has certainly led to some insights into the enzymatic structure-function relationships underlying the elongation and simpler cyclization reactions, our understanding of the more complex cyclization and/or rearrangement reactions remains limited. Accordingly, selected examples are discussed here to demonstrate our current understanding, its limits, and potential ways forward.
Summary
Danshen, Salvia miltiorrhiza Bunge, is one of the most widely used herbs in traditional Chinese medicine, wherein its rhizome/roots are particularly valued. The corresponding bioactive ...components include the tanshinone diterpenoids, the biosynthesis of which is a subject of considerable interest. Previous investigations of the S. miltiorrhiza transcriptome have relied on short‐read next‐generation sequencing (NGS) technology, and the vast majority of the resulting isotigs do not represent full‐length cDNA sequences. Moreover, these efforts have been targeted at either whole plants or hairy root cultures. Here, we demonstrate that the tanshinone pigments are produced and accumulate in the root periderm, and apply a combination of NGS and single‐molecule real‐time (SMRT) sequencing to various root tissues, particularly including the periderm, to provide a more complete view of the S. miltiorrhiza transcriptome, with further insight into tanshinone biosynthesis as well. In addition, the use of SMRT long‐read sequencing offered the ability to examine alternative splicing, which was found to occur in approximately 40% of the detected gene loci, including several involved in isoprenoid/terpenoid metabolism.
Significance Statement
Here, we combined NGS and SMRT sequencing to generate a more complete/full‐length S. miltiorrhiza transcriptome. Our study provides a general model for studying the full‐length transcriptome, alternative splicing, and the biosynthesis of active constituents in medicinal plants, as well as important information about the important Danshen constituent of traditional Chinese medicine more specifically.
Miltiradiene (1) is the precursor of phenolic diterpenoids such as ferruginol (2), requiring aromatization and hydroxylation. While this has been attributed to a single cytochrome P450 (CYP76AH1), ...characterization of the rosemary ortholog CYP76AH4 led to the discovery that these CYPs simply hydroxylate the facilely oxidized aromatic intermediate abietatriene (3).
The highly diverse structures of all terpenes are synthesized from the five carbon precursors dimethylallyl diphosphate and a varying number of isopentenyl diphosphate units through 1’-4 alkylation ...reactions. These elongation reactions are catalyzed by isoprenyl diphosphate synthases (IDS). IDS are classified depending on the configuration of the ensuing double bond as trans-and cis-IDS. In addition, IDS are further stratified by the length of their prenyl diphosphate product. This review discusses analytical techniques for the determination of product length and the factors that control product length, with an emphasis on alternative mechanisms. With recent advances in analytics, multiple IDS of Arabidopsis thaliana have been recently reinvestigated and demonstrated to yield products of different lengths than originally reported, which is summarized here. As IDS dictate prenyl diphosphate length and thereby which class of terpenes is ultimately produced, another focus of this review is the impact that altering IDS expression has on terpenoid natural product accumulation. Finally, recent findings regarding the ability of a few IDS to not catalyze 1’-4 alkylation reactions, but instead produce irregular products, with unusual connectivity, or act as terpene synthases, are also discussed.
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