Methyl transfer in psilocybin biosynthesis.

Psilocybin, the natural hallucinogen produced by Psilocybe ("magic") mushrooms, holds great promise for the treatment of depression and several other mental health conditions. The final step in the psilocybin biosynthetic pathway, dimethylation of the tryptophan-derived intermediate norbaeocystin, is catalysed by PsiM. Here we present atomic resolution (0.9 Å) crystal structures of PsiM trapped at various stages of its reaction cycle, providing detailed insight into the SAM-dependent methylation mechanism. Structural and phylogenetic analyses suggest that PsiM derives from epitranscriptomic N6-methyladenosine writers of the METTL16 family, which is further supported by the observation that bound substrates physicochemically mimic RNA. Inherent limitations of the ancestral monomethyltransferase scaffold hamper the efficiency of psilocybin assembly and leave PsiM incapable of catalysing trimethylation to aeruginascin. The results of our study will support bioengineering efforts aiming to create novel variants of psilocybin with improved therapeutic properties.

Biosynthetic incorporation of fluorinated amino acids into the nonribosomal peptide gramicidin S.

Fluorine is a key element in medicinal chemistry, as it can significantly enhance the pharmacological properties of drugs. In this study, we aimed to biosynthetically produce fluorinated analogues of the antimicrobial cyclic decapeptide gramicidin S (GS). However, our results show that the A-domain of the NRPS module GrsA rejects 4-fluorinated analogues of its native substrate Phe due to an interrupted T-shaped aromatic interaction in the binding pocket. We demonstrate that GrsA mutant W239S improves the incorporation of 4-fluorinated Phe into GS both in vitro and in vivo. Our findings provide new insights into the behavior of NRPSs towards fluorinated amino acids and strategies for the engineered biosynthesis of fluorinated peptides.