Monoamine Biosynthesis via a Noncanonical Calcium-Activatable Aromatic Amino Acid Decarboxylase in Psilocybin Mushroom

Abstract

Aromatic L-amino acid decarboxylases (AAADs) are a phylogenetically diverse group of enzymes responsible for the decarboxylation of aromatic amino acid substrates into their corresponding aromatic arylalkylamines. AAADs have been extensively studied in mammals and plants as they catalyze the first step in the production of neurotransmitters and bioactive phytochemicals, respectively. Unlike mammals and plants, the hallucinogenic psilocybin mushroom Psilocybe cubensis reportedly employs an unrelated phosphatidylserine-decarboxylase-like enzyme to catalyze Ltryptophan decarboxylation, the first step in psilocybin biosynthesis. To explore the origin of this chemistry in psilocybin mushroom, we generated the first de novo transcriptomes of P. cubensis and investigated several putative L-tryptophandecarboxylase-like enzymes. We report the biochemical characterization of a noncanonical AAAD from P. cubensis (PcncAAAD) that exhibits substrate permissiveness toward L-phenylalanine, L-tyrosine, and L-tryptophan, as well as chloro-tryptophan derivatives. The crystal structure of PcncAAAD revealed the presence of a unique C-terminal appendage domain featuring a novel double-β-barrel fold. This domain is required for PcncAAAD activity and regulates catalytic rate and thermal stability through calcium binding. PcncAAAD likely plays a role in psilocybin production in P. cubensis and offers a new tool for metabolic engineering of aromatic-amino-acid-derived natural products.

S‐Adenosyl‐l‐Methionine Salvage Impacts Psilocybin Formation in “Magic” Mushrooms

“…The trpB gene is sixfold upregulated in emerse mycelium and during mating which is consistent with an increased l ‐tryptophan demand during psilocybin biosynthesis. In contrast, the psilocybin biosynthetic gene cluster ( psiD , psiH , psiK , psiM ) was only poorly expressed in any tested mycelial stage, but its transcription was induced during fruiting body formation which is consistent with previous observations . The gene for the gateway enzyme PsiD is most upregulated (395‐fold in primordia relative to submerse mycelium).…”

Emergence of oxygen‐ and pyridoxal phosphate‐dependent reactions

“…An AAS has also been reported from Arabidopsis thaliana from its ability to convert phenylalanine and l ‐DOPA to PHA and 3,4‐dihydroxyphenylacetaldehyde (DHPAA), respectively [48]. The first known fungal AAS example, Pc DHPAAS from Psilocybe cubensis , also has activity on ʟ‐DOPA [49]. Cicer arietinum (chickpeas) and Medicago truncatula each have unique types of AASs, predicted as serine decarboxylases, rather than aromatic amino acid decarboxylases (AADCs) [50].…”

Oxygen reactivity with pyridoxal 5′-phosphate enzymes: biochemical implications and functional relevance

“…6 ), mainly related to the decarboxylation of l -tryptophan (Kalb et al 2016 ). An in-depth investigation has been carried out in Psilocybe cubensis , a mushroom responsible for psychoactive alkaloid psilocybin synthesis, revealing the presence of both a DHPAAS with a phenylalanine instead of a tyrosine in the catalytic loop and an AADC-like protein (similar to plant l -tryptophan decarboxylases) (Torrens-Spence et al 2018 ). Interestingly, in this AADC enzyme, a regulation by calcium ion was reported.…”