Crystal Structure of Codeinone Reductase Reveals Novel Features Responsible for Defining Benzylisoquinoline Alkaloid Substrate Specificity

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The pharmaceutical properties of plant-derived compounds are widely used in medicine. Benzylisoquinoline alkaloids (BIAs) are a class of specialized metabolites with a diverse range of chemical structures and physiological effects. Codeine and morphine are closely related BIAs with particularly useful analgesic properties. The aldo-keto reductase (AKR) codeinone reductase (COR) catalyzes the final step and the penultimate step in biosynthesis of codeine and morphine, respectively, in opium poppy (Papaver somniferum). Using X-ray crystallography, we have determined the structure of apo-COR (2.4 Å resolution) for the first time, using the structure of chalcone reductase as a search model. The structure of apo-COR reveals a three-dimensional framework for understanding BIA substrate recognition and catalysis. Structural comparisons of COR to closely related plant AKR's and more distantly related homologues reveal a novel conformation in the β1α1 loop adjacent to the BIA binding pocket. The proximity of this loop several highly conserved active site residues and the expected location of the nicotinamide ring of the NADPH cofactor suggest the importance of several key residues in BIA substrate recognition. The effects of mutations in residues in this loop and at other nearby positions help to define specific roles in substrate recognition and catalysis. Combined with the structure of COR, these findings from mutagenesis also help to suggest structure-function relationships in a second AKR critical to BIA biosynthesis, the 1,2 dehydroreticuline reductase domain in the critical enzyme reticuline epimerase.