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.
Taylor
M
M
AberBiocentre, Aberystwyth, Wales, SY23 3DA, United Kingdom.
Lamb
D C
DC
Cannell
R J
RJ
Dawson
M J
MJ
Kelly
S L
SL
D001426:Bacterial Proteins; D001564:Benzo(a)pyrene / Q000378:metabolism; D002474:Cell-Free System; D003001:Cloning, Molecular; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004800:Enzymes, Immobilized / Q000378:metabolism*; D004926:Escherichia coli; D007700:Kinetics; D010105:Oxygenases / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013305:Streptomyces griseus / Q000201:enzymology*; D013379:Substrate Specificity; D014859:Warfarin / Q000031:analogs & derivatives / Q000378:metabolism
D001426:Bacterial Proteins; D004800:Enzymes, Immobilized; D011994:Recombinant Proteins; D001564:Benzo(a)pyrene; D014859:Warfarin; D003577:Cytochrome P-450 Enzyme System; D010105:Oxygenases; C122191:CYP105D1 protein, Streptomyces griseus