BIOINORGANIC CHEMISTRY A Short Course Second Edition

364 IRON-CONTAINING PROTEINS AND ENZYMES

produce the ferrous species. The ferrous product is then annealed at elevated
temperatures, and the conformational and chemical relaxation processes are
studied by EPR and optical spectroscopic methods. For instance, Fe 3+ – OOH −
complexes in hemes studied by this method, including CYP101, have shown
EPRg values of 2.3 – 2.25, 2.2 – 2.14, and 1.94 – 1.97. Obtaining such EPR values
for the unstable intermediates is then taken as evidence of their presence
during the catalytic cycle. Intermediate ( 5b ) is known to dissociate to produce
hydrogen peroxide, H 2 O 2 , via the peroxide shunt upon addition of a second
proton, resulting in P450 inactivation. The next productive step in the catalytic
cycle toward the desired mono - oxygenated substrate involves addition of the
second H + to fi rst generate the unstable transient Fe – OOH 2 followed by fast
heterolytic cleavage of the O – O bond, release of H 2 O, and production of the
ferryl – oxo – porphyrin π - cation radical, Fe(IV) = O ( 6 ). This intermediate,

H 2 O 523

H 2 O 566

H 2 O 687

H 2 O 902

H 2 O 901

cys357

2.2

1.8

1.25

thr101

2.7

2.7

tyr96

3.5

2.8

3.0

3.5

2.5

3.6

1.8

2.9

2.8

3.3

thr252

asp251

asn255

glu366

PDB: 1DZ8 active site

All distances in angstroms

camphor

Figure 7.13 Cytochrome P450 catalytic site with camphor — the dioxygen intermedi-
ate (PDB: 1DZ8). Visualized using CambridgeSoft Chem3D Ultra 10.0 with notations
in ChemDraw Ultra 10.0. (Printed with permission of CambridgeSoft Corporation.)