Cofactor recycling with immobilized heterologous cytochrome P450 105D1 (CYP105D1)

Immobilisation of cells and enzymes can be a convenient and rapid way for testing and transforming substances. Cytochromes P450 may be useful in numerous biotransformations of varied lipophilic substrates, performing both regio- and stereo-specific monooxygenation reactions. However, one limitation of their use in vitro is the requirement of cofactor for the supply of electrons in the catalytic cycle. Here we report CYP105D1 from Streptomyces griseus expressed in Escherichia coli can be immobilised from cell-free extracts using DE52, that the immobilised protein is active in bioconversions and that a requirement for cofactor can be sustained by a recycling system for NADH regeneration. (C) 2000 Academic Press.