An enzymatically active chimeric protein containing the hydrophilic form of NADPH-cytochrome p450 reductase fused to the membrane-binding domain of cytochrome b(5)
Aa. Gilep et al., An enzymatically active chimeric protein containing the hydrophilic form of NADPH-cytochrome p450 reductase fused to the membrane-binding domain of cytochrome b(5), BIOC BIOP R, 284(4), 2001, pp. 937-941
Citations number
16
Categorie Soggetti
Biochemistry & Biophysics
Journal title
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
The microsomal flavoprotein NADPH-cytochrome P450 reductase (CPR) contains
an N-terminal hydrophobic membrane-binding domain required for reconstituti
on of hydroxylation activities with cytochrome P450s. In contrast, cytochro
me b(5) (b(5)) contains a C-terminal hydrophobic membrane-binding domain re
quired for interaction with P450s. We have constructed, expressed and purif
ied a chimeric flavoprotein (hdb5-CPR) where the C-terminal 45 amino acid r
esidues of b(5) have replaced the N-terminal 56 amino acid domain of CPR. T
his hybrid flavoprotein retains the catalytic properties of the native CPR
and is able to reconstitute fatty acid and steroid hydroxylation activities
with CYP4A1 and CYP17A. However hdb5-CPR is much less effective than CPR f
or reconstituting activity with CYP3A4, We conclude that differences on the
surface of the P450s reflect unique and specific information essential for
the recognition needed to establish reactions of intermolecular electron t
ransfer from the flavoprotein CPR. (C) 2001 Academic Press.