Production of human cytochrome P450 2D6 drug metabolites with recombinant microbes - a comparative study

• Published in: Biotechnology journal 2012-11, Vol.7 (11), p.1346-1358
• Main Authors: Geier, Martina, Braun, Andreas, Emmerstorfer, Anita, Pichler, Harald, Glieder, Anton
• Format: Article
• Language: English
• Subjects: Bioreactors - microbiology; Biotechnology - methods; Biotransformation; Cell Membrane; Cloning, Molecular; Cytochrome P-450 CYP2D6 - chemistry; Cytochrome P-450 CYP2D6 - genetics; Cytochrome P-450 CYP2D6 - metabolism; Escherichia coli - genetics; Escherichia coli - metabolism; Ethanolamines - metabolism; Glucose - metabolism; Host comparison; Human cytochrome P450 2D6; Humans; Membrane protein expression; NADPH-Ferrihemoprotein Reductase - analysis; NADPH-Ferrihemoprotein Reductase - chemistry; NADPH-Ferrihemoprotein Reductase - genetics; NADPH-Ferrihemoprotein Reductase - metabolism; Pichia pastoris; Plasmids; Recombinant Proteins - biosynthesis; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Saccharomycetales - genetics; Saccharomycetales - metabolism; Temperature; Whole-cell biotransformation
• ISSN: 1860-6768; 1860-7314
• DOI: 10.1002/biot.201200187

The processes of drug development require efficient strategies to produce the respective drug metabolites, which are often difficult to obtain. Biotransformations employing recombinant microorganisms as whole‐cell biocatalysts have become an attractive alternative to the chemical syntheses of such metabolites. For the first time, the potential of four different microbial systems expressing the human cytochrome P450 2D6 (CYP2D6), which is one of the most important drug‐metabolizing enzymes, were compared and evaluated for such applications. The microbial host Pichia pastoris was the most efficient at expressing CYP2D6. Without additional over‐expression of chaperons, the achieved yield of CYP2D6 was the highest of microbial hosts reported so far. Therefore, the system described in this study outperformed the previously reported expression of the N‐terminally modified enzyme. It was also shown that the activities of the whole‐cell conversions of bufuralol in recombinant P. pastoris were significantly higher than the Escherichia coli catalyst, which expressed the same unmodified gene. Biotransformations employing recombinant microorganisms as whole‐cell biocatalysts have become an attractive alternative to the chemical synthesis of drug metabolites. In this study, authors compare the potential and differences of four microbial hosts to express the human cytochrome P450 2D6 (CYP2D6). Pichia pastoris shows to be the most efficient host. Without additional over‐expression of chaperons, the achieved yield of CYP2D6 was higher than described with other microbial hosts reported so far.