River Stone Biotech LLC

Jon Richard Heal

Philipp Friedrich Berninger

Joseph Michael Sheridan

Markus Schwab

Esben Halkjaer Hansen

Franziska Grassinger

Laura Tatjer Recorda

Jens Houghton-larsen

Rubini Maya Kannangara

Sumire Honda Malca

Laura Occhipinti

Anaelle Hatsch

Angela Manuela Ribeiro De Carvalho

Esben Halkjær Hansen

Laura Tatjer RecordÁ

ÂNgela Manuela Ribeiro De Carvalho

The invention relates to norcoclaurine synthases and substrate binding sites having one or more site-specific mutation which increase the activity, when compared to the wild type synthase, of the condensation of 4-HPAA and dopamine to (S)-norcoclaurine and/or 3,4-DhPAA and dopamine to (S)-norlaudanosoline. The inventors both identified specific mutations corresponding to at position 73, 75, 77, 82, 99, 114, 141, 142, 147, 152, 174 and/or 178 in the count according to SEQ ID No: 1, and sites corresponding to the binding domains defined in SEQ ID NO: 4 and 5, where the mutated increase of the activity may be positioned within these norcoclaurine synthases. These domains are conserved regions.

Norcoclaurine synthases with increased activity

The invention relates to recombinant host cells that expresses one or more genes encoding a cytochrome P450 enzyme capable of N-demethylating and/O-demethylating reticuline and/or derivatives thereof, and also methods of producing a N-demethylated and/or O-demethylated reticuline and/or derivatives thereof, comprising cultivating the recombinant host of the invention in a culture medium under conditions in which the one or more genes encoding the cytochrome P450 enzymes is/are expressed. The reticuline and derivatives thereof are useful for providing access to naturally unavailable and chemically difficult-to-produce starting materials for opioids.

Demethylation of reticuline and derivatives thereof with fungal cytochrome P450

Growing community of inventors

Reinach, Switzerland

Jon Richard Heal