The interaction of NADPH-P450 reductase with P450: An electrochemical study of the role of the flavin mononucleotide-binding domain

Ronald W. Estabrook, Manjunath S. Shet, Charles W. Fisher, Christopher M. Jenkins, Michael R. Waterman

Research output: Contribution to journalArticlepeer-review

60 Scopus citations

Abstract

The electrochemically reduced mediator cobalt sepulchrate requires the presence of a flavoprotein for the rapid transfer of electrons to cytochrome P450. This electrochemical method has been used here to show the interaction of NADPH-P450 reductase (either the detergent-solubilized form, d-OR, or the proteolytic-cleaved truncated form, t-OR), as well as Escherichia coli flavodoxin (FLD), with P450c17 by measuring the rate of 17α-hydroxylation of progesterone. When NADPH is used as electron donor with a reconstituted system composed of d-OR and P450c17, the addition of t-OR, flavodoxin, or cytochrome c inhibited the rate of formation of 17α-hydroxyprogesterone. These results suggest the presence of a common protein binding site on the surface of d-OR, t-OR, and flavodoxin which plays a role in the interaction of the flavoproteins with the P450. It is speculated that a domain composed of acidic amino acids, located near the flavin mononucleotide-binding region of the flavoproteins, may serve as this site. No inhibition by t-OR, flavodoxin, or cytochrome c is observed when comparable experiments are carried out using the artificial recombinant fusion protein rF450[mBov17A/mRa-tOR]L1 containing the heme-domain of P450c17 linked to the flavin-domains of NADPH-P450 reductase.

Original languageEnglish
Pages (from-to)308-315
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume333
Issue number1
DOIs
StatePublished - Sep 1 1996

Fingerprint

Dive into the research topics of 'The interaction of NADPH-P450 reductase with P450: An electrochemical study of the role of the flavin mononucleotide-binding domain'. Together they form a unique fingerprint.

Cite this