Self‐Sufficient Catalytic System of Cytochrome P450cam, Putidaredoxin, and Putidaredoxin Reductase Provides a Useful Cell‐based Enzymatic Reaction

Abstract

The catalytic turnover of cytochrome P450cam (P450cam) from Pseudomonas putida requires two auxiliary reduction partners, the iron‐sulfur protein putiaredoxin (Pd) and the flavoprotein putidaredoxin reductase (PdR). We report the functional expression in Escherichia coli of tricistronic constructs consisting of P450cam encoded by the first cistron and the auxiliary proteins, Pd and PdR by the second and the third. The expression level of P450cam was 500 nmol per liter of culture. Transformed bacterial whole cells efficiently oxidized (1R)‐(+)‐camphor, the normal substrate of the enzyme, to 5‐exohydroxycamphor. More interestingly, this whole cells system successfully oxygenated a monoterpene, limonene, and converted it to (−)‐perillyl alcohol, an anticarcinogenic compound with limited availability. These bioengineered E. coli cells expressing three proteins possess a heterologous self‐sufficient P450 catalytic system that may have advantages in terms of low cost and high yield for the production of the fine chemicals. This study was supported by a Korea Research Foundation Grant funded by the Korean Government (MOEHRD, Basic Research Promotion Fund)(KRF‐2008‐331‐E00030)