Heterologous production of Cannabis sativa-derived specialised metabolites of medicinal significance – Insights into engineering strategies

Wiles, Danielle; Shanbhag, Bhuvana K.; O’Brien, Martin; Doblin, Monika S.; Bacic, Antony; Beddoe, Travis

doi:10.1016/j.phytochem.2022.113380

Cited by 10 publications

(5 citation statements)

References 182 publications

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“…Bioinformatic analyses led to the identification of two putative enzymes (Appendix B, Figure A6) that are encoded by Phatr3_J37858 and Phatr3_J2738.t1 and are capable of prenyltransferase activity, thereby allowing the production of CBGA. Both enzymes are UbiA prenyltransferase family members that could be used originally by the diatom to produce terpenes or flavonoids [49,50]. The in silico identification of THCAS and CBDAS-like endogenous candidate genes is more intricate, given the specificity of these enzymes.…”

Section: Discussionmentioning

confidence: 99%

“…Recently, cloning the Metarhizium anisopliae ARSEF23 highly reducing PKS enzyme (MaOvaA), in addition to a non-reducing PKS (MaOvaB) and a domain PKS (MaOvaC) consisting of an acyl carrier protein and a thioesterase, in the fungus Aspergillus nidulans yielded high titers of OA (80 mg/L) [55]. Thus, it would be interesting to compare the expression and behavior of cannabis enzymes regarding fungal enzymes such as highly reducing and non-reducing PKS, along with thioesterase and acyl carrier enzymes [50]. Another approach to increase OA titers could be the construction of alternative pathways using enzymes found in other OA-producing plants than cannabis, in order to achieve OA and other CB precursors while using fungal enzymes or mutation-enhanced enzymes.…”

Section: Discussionmentioning

confidence: 99%

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Bioengineering of the Marine Diatom Phaeodactylum tricornutum with Cannabis Genes Enables the Production of the Cannabinoid Precursor, Olivetolic Acid

Awwad,

Fantino,

Héneault

et al. 2023

IJMS

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The increasing demand for novel natural compounds has prompted the exploration of innovative approaches in bioengineering. This study investigates the bioengineering potential of the marine diatom Phaeodactylum tricornutum through the introduction of cannabis genes, specifically, tetraketide synthase (TKS), and olivetolic acid cyclase (OAC), for the production of the cannabinoid precursor, olivetolic acid (OA). P. tricornutum is a promising biotechnological platform due to its fast growth rate, amenability to genetic manipulation, and ability to produce valuable compounds. Through genetic engineering techniques, we successfully integrated the cannabis genes TKS and OAC into the diatom. P. tricornutum transconjugants expressing these genes showed the production of the recombinant TKS and OAC enzymes, detected via Western blot analysis, and the production of cannabinoids precursor (OA) detected using the HPLC/UV spectrum when compared to the wild-type strain. Quantitative analysis revealed significant olivetolic acid accumulation (0.6–2.6 mg/L), demonstrating the successful integration and functionality of the heterologous genes. Furthermore, the introduction of TKS and OAC genes led to the synthesis of novel molecules, potentially expanding the repertoire of bioactive compounds accessible through diatom-based biotechnology. This study demonstrates the successful bioengineering of P. tricornutum with cannabis genes, enabling the production of OA as a precursor for cannabinoid production and the synthesis of novel molecules with potential pharmaceutical applications.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Bioengineering of the Marine Diatom Phaeodactylum tricornutum with Cannabis Genes Enables the Production of the Cannabinoid Precursor, Olivetolic Acid

Awwad,

Fantino,

Héneault

et al. 2023

IJMS

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“…The production of cannabinoids and their precursors has been achieved in various heterologous hosts [49]. Complete biosynthesis of cannabinoids and their analogues was performed in Saccharomyces cerevisiae [16].…”

Section: Discussionmentioning

confidence: 99%

Metabolic Engineering of Nicotiana benthamiana to Produce Cannabinoid Precursors and Their Analogues

et al. 2022

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In recent years, the perspective towards the use of cannabis has slowly shifted from being an illicit drug to a medicinal plant. The pathway and enzymes involved in the production of cannabinoids are known; however, studies evaluating the production of cannabinoids in heterologous plants and cell cultures are still limited. In this study, we assessed the potential use of N. benthamiana (Nicotiana benthamiana) plants as a heterologous host for producing natural and novel cannabinoids. Transgenic N. benthamiana plants expressing genes encoding cannabis acyl-activating enzyme and olivetol synthase were generated, which were then used for transiently expressing other downstream pathway genes. Production of olivetolic acid and divarinic acid, the universal precursors for major and minor cannabinoids, respectively, was observed in transgenic N. benthamiana plants. To produce novel cannabinoid precursors with different side chains, various fatty acids were infiltrated into the transgenic N. benthamiana plants and the production of novel derivatives was observed. Although we were not able to derive the core intermediate, cannabigerolic acid, from our transgenic plants, possibly due to the low production levels of the precursors, our transgenics plants still serve as a high-potential platform for further development and exploring the N. benthamiana chemical space for generating novel cannabinoids.

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“…For instance, the accumulation of cannflavin A is predisposed to genetic variations, as well as environmental factors such as temperature, rainfall, and humidity in the environment ( Kumar and Pandey, 2013 ). Besides, the contents of cannflavin A, B, and C in cloned species of C. sativa vary at different altitudes ( Wiles et al, 2022 ). With these findings, it can be postulated that identifying unknown flavonoids in the plant, is reliant on certain environmental conditions or stresses.…”

Section: Phytochemistry Of Cannabismentioning

confidence: 99%

Cannabis: a multifaceted plant with endless potentials

et al. 2023

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Cannabis sativa, also known as “hemp” or “weed,” is a versatile plant with various uses in medicine, agriculture, food, and cosmetics. This review attempts to evaluate the available literature on the ecology, chemical composition, phytochemistry, pharmacology, traditional uses, industrial uses, and toxicology of Cannabis sativa. So far, 566 chemical compounds have been isolated from Cannabis, including 125 cannabinoids and 198 non-cannabinoids. The psychoactive and physiologically active part of the plant is a cannabinoid, mostly found in the flowers, but also present in smaller amounts in the leaves, stems, and seeds. Of all phytochemicals, terpenes form the largest composition in the plant. Pharmacological evidence reveals that the plants contain cannabinoids which exhibit potential as antioxidants, antibacterial agents, anticancer agents, and anti-inflammatory agents. Furthermore, the compounds in the plants have reported applications in the food and cosmetic industries. Significantly, Cannabis cultivation has a minimal negative impact on the environment in terms of cultivation. Most of the studies focused on the chemical make-up, phytochemistry, and pharmacological effects, but not much is known about the toxic effects. Overall, the Cannabis plant has enormous potential for biological and industrial uses, as well as traditional and other medicinal uses. However, further research is necessary to fully understand and explore the uses and beneficial properties of Cannabis sativa.

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Heterologous production of Cannabis sativa-derived specialised metabolites of medicinal significance – Insights into engineering strategies

Cited by 10 publications

References 182 publications

Bioengineering of the Marine Diatom Phaeodactylum tricornutum with Cannabis Genes Enables the Production of the Cannabinoid Precursor, Olivetolic Acid

Bioengineering of the Marine Diatom Phaeodactylum tricornutum with Cannabis Genes Enables the Production of the Cannabinoid Precursor, Olivetolic Acid

Metabolic Engineering of Nicotiana benthamiana to Produce Cannabinoid Precursors and Their Analogues

Cannabis: a multifaceted plant with endless potentials

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