Biotransformation, Using Recombinant CYP450-Expressing Baker’s Yeast Cells, Identifies a Novel CYP2D6.10A122V Variant Which Is a Superior Metabolizer of Codeine to Morphine Than the Wild-Type Enzyme
Authors: Ibidapo S Williams
Sandip B Bharate
Linda Gatchie
Bhabatosh Chaudhuri
Journal: ACS Omega (2018)
Discipline: CHEMISTRY
Keywords: CYP2D6; codeine; morphine; genetic polymorphisms; yeast cells
Affiliations: CYP Design Ltd, The Innovation Centre, 49 Oxford Street, Leicester LE1 5XY, U.K., Medicinal Chemistry Division, CSIR-Indian Institute of Integrative Medicine, Canal Road, Jammu 180001, India

Abstract:  CYP2D6, a cytochrome P450 (CYP) enzyme, metabolizes codeine to morphine. Within the human body, 0-15% of codeine undergoes O-demethylation by CYP2D6 to form morphine, a far stronger analgesic than codeine. Genetic polymorphisms in wild-type CYP2D6 (CYP2D6-wt) are known to cause poor-to-extensive metabolism of codeine and other CYP2D6 substrates. We have established a platform technology that allows stable expression of human CYP genes from chromosomal loci of baker's yeast cells. Four CYP2D6 alleles, (i) chemically synthesized CYP2D6.1, (ii) chemically synthesized CYP2D6-wt, (iii) chemically synthesized CYP2D6.10, and (iv) a novel CYP2D6.10 variant CYP2D6-C (i.e., CYP2D6.10A122V) isolated from a liver cDNA library, were cloned for chromosomal integration in yeast cells. When expressed in yeast, CYP2D6.10 enzyme shows weak activity compared with CYP2D6-wt and CYP2D6.1 which have moderate activity, as reported earlier. Surprisingly, however, the CYP2D6-C enzyme is far more active than CYP2D6.10. More surprisingly, although CYP2D6.10 is a known low metabolizer of codeine, yeast cells expressing CYP2D6-C transform &gt;70% of codeine to morphine, which is more than twice that of cells expressing the extensive metabolizers, CYP2D6.1, and CYP2D6-wt. The latter two enzymes predominantly catalyze formation of codeine's N-demethylation product, norcodeine, with &gt;55% yield. Molecular modeling studies explain the specificity of CYP2D6-C for O-demethylation, validating observed experimental results. The yeast-based CYP2D6 expression systems, described here, could find generic use in CYP2D6-mediated drug metabolism and also in high-yield chemical reactions that allow the formation of regio-specific dealkylation products. 

Retracted: No

PubPeer comments:
 Would the authors be able to post the uncropped blot supporting figure 3A? *** Inverse image ![file] *** ![file] , 
 #1 Just curious, how can you see that the image was cropped? , 
 #1 Dear Pupbeer, I understand your concern over the western blot picture in this paper. Unfortunately, the original western blot data is in the university (DMU Leicester). The corresponding author Professor Bhabatosh Chaudhuri has untimely retired due to his health deterioration. Thus, the data cannot be accessed anymore. All data I have with me is the data that I generated at the time of the experiment which is placed in a PowerPoint file. I assure you that the data presented in the manuscript is completely original. As a lead biology author, I state that the data and results presented are true output of the actually done Western-blot experiment. I hope this would satisfy your query. Best wishes, Ibidapo , 
 The comments pertaining to this publication are related to the biological experiments (Western-blots) which were conducted by authors, Ibidapo S. Williams, Linda Gatchie, and mentored by Prof. Bhabatosh Chaudhuri at CYP Design Ltd, UK. My group only performed the molecular modeling studies (shown in Fig. 5) in this publication. Thus, there are no clarifications against these comments from my side. At my end, these authors have been contacted via email. One of the biology authors, Ibidapo S. Williams has already responded at this forum. Further clarifications (if any) will be provided by the biology authors mentioned above. ,