Evolution of catalytic microenvironment governs substrate and product diversity in trichodiene synthase and other terpene fold enzymes

Kumari, Indu; Ahmed, Mushtaq; Akhter, Yusuf

doi:10.1016/j.biochi.2017.10.003

Cited by 5 publications

(5 citation statements)

References 63 publications

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“…IPP is further isomerized to DMAPP by the enzyme IPP isomerase. GPP is the precursor to monoterpenes, and their biosynthesis is mediated by terpene synthases [6][7][8]. Within heterobranchs, in this time period, only anaspideans and sacoglossans are found to possess monoterpenoids, which may be linear or cyclic (Table 1).…”

Section: Monoterpenoidsmentioning

confidence: 99%

Terpenoids in Marine Heterobranch Molluscs

Ávila

2020

Marine Drugs

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Heterobranch molluscs are rich in natural products. As other marine organisms, these gastropods are still quite unexplored, but they provide a stunning arsenal of compounds with interesting activities. Among their natural products, terpenoids are particularly abundant and diverse, including monoterpenoids, sesquiterpenoids, diterpenoids, sesterterpenoids, triterpenoids, tetraterpenoids, and steroids. This review evaluates the different kinds of terpenoids found in heterobranchs and reports on their bioactivity. It includes more than 330 metabolites isolated from ca. 70 species of heterobranchs. The monoterpenoids reported may be linear or monocyclic, while sesquiterpenoids may include linear, monocyclic, bicyclic, or tricyclic molecules. Diterpenoids in heterobranchs may include linear, monocyclic, bicyclic, tricyclic, or tetracyclic compounds. Sesterterpenoids, instead, are linear, bicyclic, or tetracyclic. Triterpenoids, tetraterpenoids, and steroids are not as abundant as the previously mentioned types. Within heterobranch molluscs, no terpenoids have been described in this period in tylodinoideans, cephalaspideans, or pteropods, and most terpenoids have been found in nudibranchs, anaspideans, and sacoglossans, with very few compounds in pleurobranchoideans and pulmonates. Monoterpenoids are present mostly in anaspidea, and less abundant in sacoglossa. Nudibranchs are especially rich in sesquiterpenes, which are also present in anaspidea, and in less numbers in sacoglossa and pulmonata. Diterpenoids are also very abundant in nudibranchs, present also in anaspidea, and scarce in pleurobranchoidea, sacoglossa, and pulmonata. Sesterterpenoids are only found in nudibranchia, while triterpenoids, carotenoids, and steroids are only reported for nudibranchia, pleurobranchoidea, and anaspidea. Many of these compounds are obtained from their diet, while others are biotransformed, or de novo biosynthesized by the molluscs. Overall, a huge variety of structures is found, indicating that chemodiversity correlates to the amazing biodiversity of this fascinating group of molluscs.

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Section: Monoterpenoidsmentioning

confidence: 99%

Terpenoids in Marine Heterobranch Molluscs

Ávila

2020

Marine Drugs

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“…This result was also supported by structural studies of other plant terpene synthases 5 , 9 , 25 , 72 . It was reported that arginine, phenylalanine, tyrosine, valine, tryptophan and isoleucine were the commonly observed amino acid residues at the catalytic site of the terpene synthases 87 , which was also observed in the Pam Tps1 active site. The presence of aromatic residue pairs (Y523 and W268) at the bottom of the active site did not appear to restrict the size of the active site, and the hydrocarbon group of FPP appeared to fit perfectly into the catalytic pocket, which may shed light on the possibility of Pam Tps1 accepting FPP as a substrate (Fig.…”

Section: Resultsmentioning

confidence: 92%

Kinetic studies and homology modeling of a dual-substrate linalool/nerolidol synthase from Plectranthus amboinicus

Ashaari

Mohd-Hairul

Sabri

et al. 2021

Sci Rep

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Linalool and nerolidol are terpene alcohols that occur naturally in many aromatic plants and are commonly used in food and cosmetic industries as flavors and fragrances. In plants, linalool and nerolidol are biosynthesized as a result of respective linalool synthase and nerolidol synthase, or a single linalool/nerolidol synthase. In our previous work, we have isolated a linalool/nerolidol synthase (designated as PamTps1) from a local herbal plant, Plectranthus amboinicus, and successfully demonstrated the production of linalool and nerolidol in an Escherichia coli system. In this work, the biochemical properties of PamTps1 were analyzed, and its 3D homology model with the docking positions of its substrates, geranyl pyrophosphate (C10) and farnesyl pyrophosphate (C15) in the active site were constructed. PamTps1 exhibited the highest enzymatic activity at an optimal pH and temperature of 6.5 and 30 °C, respectively, and in the presence of 20 mM magnesium as a cofactor. The Michaelis–Menten constant (Km) and catalytic efficiency (kcat/Km) values of 16.72 ± 1.32 µM and 9.57 × 10–3 µM−1 s−1, respectively, showed that PamTps1 had a higher binding affinity and specificity for GPP instead of FPP as expected for a monoterpene synthase. The PamTps1 exhibits feature of a class I terpene synthase fold that made up of α-helices architecture with N-terminal domain and catalytic C-terminal domain. Nine aromatic residues (W268, Y272, Y299, F371, Y378, Y379, F447, Y517 and Y523) outlined the hydrophobic walls of the active site cavity, whilst residues from the RRx8W motif, RxR motif, H-α1 and J-K loops formed the active site lid that shielded the highly reactive carbocationic intermediates from the solvents. The dual substrates use by PamTps1 was hypothesized to be possible due to the architecture and residues lining the catalytic site that can accommodate larger substrate (FPP) as demonstrated by the protein modelling and docking analysis. This model serves as a first glimpse into the structural insights of the PamTps1 catalytic active site as a multi-substrate linalool/nerolidol synthase.

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“…Salt–bridge interactions are formed of the pyrophosphate moiety with the neighboring charged residues, such as R305, while hydrogen-bond networks exist with polar residues, such as N219, S223, and Y315 ( Figure 4 B). At the bottom of the binding pocket, nine residues (F57, F76, F77, Y146, I177, V179, T182, N215, and W308) directly participate in forming the catalytic domain of PentS [ 12 , 37 ]. For the benzene rings of F57, F76, and F77 and indole ring of W308, all of them make hydrophobic contact with the prenyl tail of FPP ( Figure 4 C).…”

Section: Resultsmentioning

confidence: 99%

A Single Active-Site Mutagenesis Confers Enhanced Activity and/or Changed Product Distribution to a Pentalenene Synthase from Streptomyces sp. PSKA01

et al. 2023

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Pentalenene is a ternary cyclic sesquiterpene formed via the ionization and cyclization of farnesyl pyrophosphate (FPP), which is catalyzed by pentalenene synthase (PentS). To better understand the cyclization reactions, it is necessary to identify more key sites and elucidate their roles in terms of catalytic activity and product specificity control. Previous studies primarily relied on the crystal structure of PentS to analyze and verify critical active sites in the active cavity, while this study started with the function of PentS and screened a novel key site through random mutagenesis. In this study, we constructed a pentalenene synthetic pathway in E. coli BL21(DE3) and generated PentS variants with random mutations to construct a mutant library. A mutant, PentS-13, with a varied product diversity, was obtained through shake-flask fermentation and product identification. After sequencing and the functional verification of the mutation sites, it was found that T182A, located in the G2 helix, was responsible for the phenotype of PentS-13. The site-saturation mutagenesis of T182 demonstrated that mutations at this site not only affected the solubility and activity of the enzyme but also affected the specificity of the product. The other products were generated through different routes and via different carbocation intermediates, indicating that the 182 active site is crucial for PentS to stabilize and guide the regioselectivity of carbocations. Molecular docking and molecular dynamics simulations suggested that these mutations may induce changes in the shape and volume of the active cavity and disturb hydrophobic/polar interactions that were sufficient to reposition reactive intermediates for alternative reaction pathways. This article provides rational explanations for these findings, which may generally allow for the protein engineering of other terpene synthases to improve their catalytic efficiency or modify their specificities.

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Evolution of catalytic microenvironment governs substrate and product diversity in trichodiene synthase and other terpene fold enzymes

Cited by 5 publications

References 63 publications

Terpenoids in Marine Heterobranch Molluscs

Terpenoids in Marine Heterobranch Molluscs

Kinetic studies and homology modeling of a dual-substrate linalool/nerolidol synthase from Plectranthus amboinicus

A Single Active-Site Mutagenesis Confers Enhanced Activity and/or Changed Product Distribution to a Pentalenene Synthase from Streptomyces sp. PSKA01

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