Genetically engineered bacterial cells co-expressing human cytochrome P450 with NADPH-cytochrome P450 reductase: prediction of metabolism and toxicity of drugs in humans.
Genetically engineered bacterial cells expressing human cytochrome P450 (CYP) have been developed as new tools to predict the metabolism and toxicity of drugs in humans. There are various host cells for the heterologous expression of a form of CYP. Among them, bacterial cells such as Escherichia coli (E. coli) have advantages with regard to ease of use and high yield of protein. CYP protein could be first expressed by the modification of the N-terminal amino acid sequence in E. coli cells in 1991. Since then, many forms of human CYP have been successfully expressed in E. coli cells. Since the E. coli cells do not possess endogeneous electron transport systems to support the full catalytic activity of CYP, E. coli strains co-expressing both human CYP and NADPH-cytochrome P450 reductase (OR) have been established. Each form of CYP expressed in the E. coli cells efficiently catalyzed the oxidation of a representative substrate at an efficient rate, indicating that the OR was sufficiently expressed to support the catalytic activity of CYP. According to the studies performed so far, the modification of the N-terminal amino acid sequence of CYP did not seem to affect the catalytic properties of CYP. The human CYP expressed in the E. coli cells were applicable for studies to determine a metabolic pathway(s) of drugs and to estimate kinetic parameters of drug metabolism by human CYP. Drug-drug interactions caused by inhibition of the metabolism of drugs by human CYP could also be examined by in vitro inhibition studies with CYP expressed in the E. coli cells. Recently, human CYP was co-expressed with the OR in Salmonella typhimurium (S. typhimurium) cells used for mutation assay (Ames test) by applying the technology for the expression of human CYP and the OR in E. coli cells, to evaluate whether chemicals including drugs are metabolically activated by human CYP and show mutagenicity. These strains of bacteria are considered as useful tools to study the metabolism and the toxicity of drugs in humans.
Fujita
Ken-Ichi
K
Laboratory of Drug Metabolism, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo, Japan. fujita@pharm.hokudai.ac.jp
Kamataki
Tetsuya
T