CYTOCHROME P450 : A MONOOXYGENASE 373
Fe III with the cysteine - like ligand, β - mercaptoethanol (BME) thiolate, an
anionic sulfur donor, residing in the proximal cavity. Because of glycine ’ s
smaller size, the H93G cavity mutant has a larger proximal pocket that will
accommodate foreign natural ligands like BME. This cysteinate - ligated heme
complex serves as a structural model for cytochrome P450 as well as other
enzymes with similar cofactor cores such asCalderiomyces fumago chloroper-
oxidase and nitric oxide synthase. It is known that cysteine must be deproton-
ated to cysteinate throughout the catalytic cycles of these enzymes. Dawson
has postulated that a “ push ” from the deprotonated cysteine ligand is impor-
tant for the monooxygenase activity of cytochrome P450.^59 It is known that
three amide N – H · · · S hydrogen bonds from leu358, gly359, and gln360 help
retain cysteinate coordination in the proximal pocket throughout the catalytic
cycle as iron changes its oxidation from Fe II to Fe III to Fe IV and back again. In
the PDB: 1EVP crystal structure, the BME ligand appears to have an intra-
molecular hydrogen bond that mimics one of the necessary N – H · · · S hydrogen
bonds in cytochrome P450. The reference 59 authors compare the structure of
the model complex, ferric H93G - BME Mb, with the Schlichting X - ray crystal-
lographic study of the analogous fi ve - coordinate, high - spin Fe III P450 CAM
enzyme, PDB: 1DZ4. This structure, which has two symmetry - independent
molecules, is discussed in Section 7.4.4 above. Comparison of bond distances
and angles are collected in Table 7.3.
Note that the S γ – Fe III – C bond angle for the cysteinate ligand suggests that
the sulfur atom issp^3 hybridized and that the Fe III – S bond is covalent. The
water molecule at 4.2 Å distance from Fe III in the distal pocket is within hydro-
gen bonding distance to distal his64 (3.0 Å ) The good agreement in the struc-
tural data shows that Fe III H93G - BME Mb is an excellent protein - based model
system for Fe III P450 CAM. Spectral data are in good agreement as well. The
UV – vis spectrum of Fe III H93G - BME Mb shows a prominent peak at 618 nm
that is typical of high - spin ferric heme complexes, and its magnetic circular
TABLE 7.3 Bond Distances and Angles for Fe III H93G - BME Mb with Fe III
P450CAM
Bond Distance ( Å ) or Bond
Angle ( ° ) Fe III H93G - BME Mb Fe III P450 CAMa
Proximal ligand, Fe III – S 2.41 2.37
2.36
Distal ligand None closer than 4 Å None closer than 4 Å
H 2 O at 4.22 Å
Distance between Fe III and
porphyrin plane
0.4 0.4S γ – Fe III – C bond angle 111 108
110
a Data given for two symmetry - independent molecules in the unit cell.