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For the characterization of the substrate binding site optical and EPR measurements with spin labelled substrates on solubilized and pure cytochrome P-450 were performed. Analogously to the unlabelled derivatives spin labelled n-alkylamines and isocyanides with different chain lengths are type II substrates. The Ks-values evaluated from optical (P-450 = 1.98 . 10(-6) M) and ESR (P-450 = 1.98 . 10(-4) M) measurements are very similar indicating no concentration dependences. Contrary to the unlabelled n-alkylamines the spin labelled compounds show an affinity almost independent of the chain lengths. The SL-substrates with a short distance between the functional group and the NO-group bound to P-450 induce pronounced changes of the ligand field of the heme iron and a large broadening of the signal of the immobilized nitroxide indicating intensive interactions between the unpaired electron of the nitroxide group and the paramagnetic heme iron. Elongation of the alkyl chains results in spectra of the Fe3+ complexes with only slight modification and a remained unbroadened signal of the immobilized nitroxide. The binding of the substrate through their functional groups together with a 1:1 stoichiometry of the P-450 SL-IC-complex give evidence for the same binding site in the near vicinity of the heme iron.
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
November 1977,
FEBS letters,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
February 1984,
Biochemical and biophysical research communications,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
January 1979,
Acta biologica et medica Germanica,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
June 1983,
Biokhimiia (Moscow, Russia),
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
April 1986,
Archives of biochemistry and biophysics,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
June 1978,
The Journal of biological chemistry,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
July 1989,
Mutation research,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
March 1987,
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J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
June 1976,
European journal of biochemistry,
J Pirrwitz, and
G Lassmann, and
H Rein, and
G R Jänig, and
S Pecar, and
K Ruckpaul
July 1983,
The Journal of biological chemistry,
