Psilocybin, the psychoactive tryptamine-derived natural product from Psilocybe mushrooms, has had its biosynthetic enzymes characterized. Four enzymes were identified: PsiD, a new class of fungal L-tryptophan decarboxylase; PsiK, catalyzing the phosphotransfer step; PsiM, a methyltransferase performing repeated N-methylation as the final step; and PsiH, a monooxygenase. A step-economical route synthesized psilocybin from 4-hydroxy-L-tryptophan using PsiD, PsiK, and PsiM in a combined reaction. These findings may enable biotechnological production, given renewed pharmaceutical interest.
Psilocybin, the main psychoactive alkaloid in magic mushrooms, is produced by several fungal genera. In Psilocybe species the biosynthetic pathway from L-tryptophan is well characterized. This work examined the pathway in Inocybe corydalina, a distantly related mushroom that also makes psilocybin. Four recombinant enzymes from I. corydalina were characterized in vitro: a decarboxylase (IpsD), a kinase (IpsK), and two methyltransferases (IpsM1 and IpsM2); a fifth monooxygenase (IpsH) was analyzed in silico. None of the reactions matched those in Psilocybe. The Inocybe pathway is branched and also produces baeocystin as a second end product. The findings indicate that psilocybin biosynthesis evolved twice independently using unrelated enzymes.