Using biotinylated adrenodoxin and avidin-Sepharose 4B, dissociation constants for the complex between adrenodoxin reductase and adrenodoxin in the oxidized and reduced states were determined as 50 +/- 11 and 296 +/- 44 nM, respectively. Concentrations of adrenodoxin reductase/adrenodoxin in the matrix fraction from bovine adrenal cortex, liver, and kidney mitochondria were determined to be 20.2 +/- 10.6 mu M/ 120 +/- 23 mu M, 0.17 +/- 0.06 mu M/1.79 +/- 0.24 mu M, and 0.40 +/- 0.23 mu M/1.33 +/- 0.26 mu M, respectively. The calculation of the percentage of adrenodoxin reductase in the complex form in the reduced state showed that it is higher than 99% in adrenal cortex mitochondria, providing clear evidence for the cluster model for the mitochondrial steroid hydroxylase system. (C) 2000 Academic Press.
