Biosynthesis and synthetic biology of psychoactive natural products

Abstract

Psychoactive natural products play an integral role in the modern world. The tremendous structural complexity displayed by such molecules confers diverse biological activities of significant medicinal value and sociocultural impact. Accordingly, in the last two centuries, immense effort has been devoted towards establishing how plants, animals, and fungi synthesize complex natural products from simple metabolic precursors. The recent explosion of genomics data and molecular biology tools has enabled the identification of genes encoding proteins that catalyze individual biosynthetic steps. Once fully elucidated, the "biosynthetic pathways" are often comparable to organic syntheses in elegance and yield. Additionally, the discovery of biosynthetic enzymes provides powerful catalysts which may be repurposed for synthetic biology applications, or implemented with chemoenzymatic synthetic approaches. In this review, we discuss the progress that has been made toward biosynthetic pathway elucidation amongst four classes of psychoactive natural products: hallucinogens, stimulants, cannabinoids, and opioids. Compounds of diverse biosynthetic origin - terpene, amino acid, polyketide - are identified, and notable mechanisms of key scaffold transforming steps are highlighted. We also provide a description of subsequent applications of the biosynthetic machinery, with an emphasis placed on the synthetic biology and metabolic engineering strategies enabling heterologous production.

Fig. 1.

Four categories of psychoactive natural products or derivatives described in this review.

Fig. 2:. PLP-Dependent amino acid decarboxylase.

(A) three amino acids are decarboxylated to give primary amines that are building blocks for alkaloids; (B) mechanism of the PLP-dependent tryptophan decarboxylase

Fig. 3:. Mannich reactions in alkaloid biosynthesis.

(A) formation of the pyrrolidine intermediate on pathway to tropane alkaloids; (B) the Pictet-Spengler reaction involving tryptamine to form tetrahydro-β-carboline intermediates; (C) the Pictet-Spengler reaction involving dopamine to form tetrahydroisoquinoline on pathway to morphine.

Fig. 4:. Enzyme catalyzed group transfer reactions in biosynthesis.

(A) acetyltransferase-catalyzed acetyltransfer; (B) methyltransferase-catalyzed methyl transfer; (C) glucosyltransferase-catalyzed glucosyl transfer.; and (D) prenyltransferase-catalyzed prenyl transfer.

Fig. 5:. Two examples of P450 catalyzed oxidative modifications in biosynthesis of plant natural products.

(A) secologanin synthase in biosynthesis of monoterpene indole alkaloids; (B) salutaridine synthase in biosynthesis of morphine family of opioids.

Fig. 6.

Strategies in synthetic biology.

Fig. 7.

Amino acid building blocks for hallucinogens that target serotonin receptors.

Fig. 8.. Overview of hallucinogenic natural products.

*Note that LSD 3 is a semisynthetic compound derived from lysergic acid (Section 2.5).

Fig. 9.. Psychotria viridis is one of the common sources of DMT for ritual purposes.

Image on the left courtesy of Paulo Pedro P. R. Costa via. CC-4.0. https://upload.wikimedia.org/wikipedia/commons/0/01/PsychotriaviridisFrutoDSC75.jpg

Fig. 10.. Incilius alvarius’s skin and exudates contain 5-methoxy-N,N-ditryptamine and bufotenin.

Image on top courtesy of Wildfeurer via. CC-3.0. https://upload.wikimedia.org/wikipedia/commons/4/4f/2009-03-13Bufo_alvarius067.jpg

Fig. 11.

Biosynthesis of DMT.

Fig. 12.. Psilocybe mexicana contains ~1% psilocybin.

Image on left courtesy of Alan Rockefeller via CC-3.0. https://upload.wikimedia.org/wikipedia/commons/4/46/Psilocybe_mexicana_53960.jpg

Fig. 13.

Biosynthetic pathway of psilocybin and psilocin from l-tryptophan.

Fig. 14.

Engineered production of psilocybin in E. coli.

Fig. 15.

Engineered production of psilocybin and psilocin in yeast.

Fig. 16.. Banisteriopsis caapi contains many compounds with the β-carboline scaffold, including harmine.

Image on left courtesy Forest and Kim Starr via CC-2.0. https://upload.wikimedia.org/wikipedia/commons/1/17/Starr-140222-0335-Banisteriopsis_caapi-leaves-Haiku-Maui_%2825240510635%29.jpg

Fig. 17.

Proposed biosynthesis of harmala alkaloids.

Fig. 18.. Claviceps purpurea (ergot fungus) infecting Dactylis glomerata (cat grass).

Image on the left courtesy of Bildoj via CC-3.0. https://upload.wikimedia.org/wikipedia/commons/c/c4/Dactylis_026.JPG

Fig. 19.

Biosynthesis of lysergic acid from l-tryptophan.

Fig. 20.. Lophophora williamsii, one of the many cacti species that contain mescaline.

Image on the left courtesy of Peter A. Mansfeld via CC-3.0. https://upload.wikimedia.org/wikipedia/commons/6/69/Lophophora_williamsii_pm.jpg

Fig. 21.

Proposed biosynthesis of mescaline.

Fig. 22.

Amanita muscaria contains about ~100–1000 ppm of ibotenic acid and muscimol.

Fig. 23.

Biosynthesis of ibotenic acid and muscimol from l-glutamic acid.

Fig. 24.. Tabernanthe iboga in fruit.

Image courtesy of Christian Kunath via CC-3.0. https://twitter.com/sesamothamnus/status/1031998713760231424

Fig. 25.

Biosynthesis of secologanin from geranyl pyrophosphate (GPP).

Fig. 26.

Biosynthesis of ibogaine from tryptamine and secologanin.

Fig. 27.

Heterologous production of strictosidine in S. cerevisiae.

Fig. 28.

Salvia divinorum contains salvinorin A, a structurally unique terpene hallucinogen. Image on the left courtesy of Eric Hunt via CC-2.5. https://upload.wikimedia.org/wikipedia/commons/3/35/Salvia_divinorum_-1.jpg

Fig. 29.

Proposed biosynthetic pathway for salvinorin A.

Fig. 30.

Alkaloidal stimulants as structural mimics of neurotransmitters.

Fig. 31.

Coffea arabica (the dominant coffee cultivar) contains ~1.2 percent dry weight caffeine.

Fig. 32.. Caffeine biosynthesis and microbial engineering strategies.

(A) Major caffeine biosynthetic route identified in Camellia sinensis and Coffea arabica. (B) SAH-derived adenosine may be funneled into purine metabolism in tea leaves following methyl transfer. (C) Xanthine recycle pathway utilized during heterologous production in yeast. (D) Novel xanthine-to-caffeine conversion pathway leveraged for caffeine production in E. coli.

Fig. 33.

Nicotiana tabacum leaves contain 2 to 8 percent dry weight nicotine.

Fig. 34.. Summary of the nicotine biosynthetic pathway, including known and proposed enzymatic steps.

(A) N-methylpyrrolinium formation via the polyamine pathway. (B) Proposed reduction of nicotinic acid via A622. (C) Proposed oxidation of condensation products via BBL towards nicotine, nornicotine.

Fig. 35.. Erythroxylum coca leaves contain ~0.7 percent dry weight cocaine.

Image on left courtesy of Danna Lizeth Guevara Prieto via CC-4.0. https://www.inaturalist.org/photos/22483426

Fig. 36.

Formation of tropine, pseudotropine.

Fig. 37.

Scopolamine biosynthesis from phenylalanine and tropine.

Fig. 38.. Cocaine biosynthesis.

(A) Racemization of the cocaine pathway intermediate decarboxylation product hygrine. (B) Proposed biosynthesis of methylecognone and subsequent formation of cocaine.

Fig. 39.

Production of tropane alkaloids in yeast.

Fig. 40.

The Cannabis sativa plant typically contains 5–16% tetrahydrocannabinol (7).

Fig. 41.

Structural motifs and examples of isolated natural products from the Cannabis plant.

Fig. 42.. Exemplary structurally related cannabinoid-like natural products isolated from other plant and fungal sources (italics).

Structural deviations highlighted in red. Amorfrutin 2 (B) (148) is a 148 derivative, (–)-cis-perrottetinene (149) is a 7 derivative, machaeridiol (150) is a 8 derivative, and 6-chloro-cannabiorchichromene (151) is a 144 derivative.

Fig. 43.

CB1 and CB2 activity for 7, 8, 147, the natural endocannabinoid arachidonylcyclopropylamide, and synthetic analogue JWH-133.

Fig. 44.

CB1 and CB2 activity of THC (7) with varying C3 alkyl chain lengths, propyl (varin, 152) and heptyl (phorol, 154). CBD alkyl chain length derivatives also shown for clarity.

Fig. 45.

Biosynthesis of cannabigerol (147) and cannabinerol (159) from hexanoyl-CoA 156 and malonyl-CoA 127.

Fig. 46.

Biosynthesis of tetrahydrocannabinol (7), cannabidiol (8), cannabichromene (144), and further nonenzymatic derivatized products.

Fig. 47.. Key proposed step in biosynthesis of cannabis natural products converting cannabigerolic or cannabinerolic acid to THCA (160).

(A) Enzymatic dehydrogenation reaction leads to a reactive quinone methide intermediate 166 that can undergo various pericyclic reactions to yield all cannabis scaffolds. Flavin adenine dinucleotide (FAD), R = C₁₆H₂₆N₅O₁₃P₂. (B) Related enzymatic transformations by CBCAS and CBDAS form CBCA (162) and CBDA (161)

Fig. 48.

Heterologous production of tetrahydrocannabinolic acid (160) and cannabidiolic acid (161).

Fig 49.

Cell-free system for improved olivetolic acid, divarinic acid, and geranyl pyrophosphate production.

Fig. 50.. Image of bulbs and bloom of the poppy plant, Papaver somniferum.

On average, poppy bulbs contain 16% by weight morphine 9.

Fig. 51.

Structures of natural morphinan opioids and synthetic compound diacetylmorphine (heroin, 169).

Fig. 52.

Structures of simple benzyl isoquinolines that play key roles in opioid biosynthesis (172, 28, 27) and as antispasmodic drugs (173).

Fig. 53.

Phthalide isoquinoline opioid natural products, noscapine (174) and narceine (175).

Fig. 54.

Aporphine opioids corytuberine (176), natural (S)-glaucine (177) and unnatural (R)-glaucine (178).

Fig. 55.

Examples of berberine opioids, berberine (179) and sanguinarine (180).

Fig. 56.

Biosynthesis of the morphine opioids from dopamine 17 and 4-hydroxyphenylacetaldehyde 26.

Fig. 57.

Noscapine biosynthesis from (S)-reticuline.

Fig. 58.

Heterologous production of thebaine and hydrocodone from sugar in yeast.

Fig. 59.. Mitragyna speciosa cultivars may contain up to one percent dry weight mitragynine.

Image on left courtesy of Thor Porre via CC-3.0. https://commons.wikimedia.org/wiki/File:Kratom_tree.jpg

Fig. 60.

Proposed biosynthetic route to kratom alkaloids.