In Vitro Psilocybin Synthesis by Co‐Immobilized Enzymes

Advanced clinical trials investigate the Psilocybe magic mushroom natural product psilocybin as a treatment against major depressive disorder. Currently, synthetic material is used to meet the demand for legitimate pharmaceutical purposes. Here, we report an in vitro approach to biocatalytically produce psilocybin on a solid-phase matrix charged with five covalently bound biosynthetic enzymes. These enzymes include three Psilocybe enzymes: IasA*, an engineered l-tryptophan decarboxylase/aromatic aldehyde synthase, the 4-hydroxytryptamine kinase PsiK and the norbaeocystin methyltransferase PsiM, along with Escherichia coli nucleosidase MtnN and adenine deaminase Ade. In a proof-of-principle experiment, this enzyme-charged resin allowed for quantitative turnover of 4-hydroxy-l-tryptophan into psilocybin. This facile process i) represents a sustainable approach with reusable enzymes, ii) circumvents the drawbacks of in vivo processes while harnessing the selectivity of enzymatic catalysis and iii) helps access an urgently needed drug candidate.