CYTOCHROMES c 413
electronegative “ heavy ” atoms in a hydrogen - bonded pair ( dAD ) must be less
than 3.4 Å ; and (3) the D – H – A angle must be greater than 90 °.^111 Since most
X - ray crystallographic structures do not display hydrogen atoms, criteria (2)
will be used most frequently in this work. The protoheme IX group numbering
system, as outlined in reference 112 , is shown in Figure 7.32B.
The PDB: 2YCC X - ray crystallographic structure, with Fe(III) in the heme,
refi ned to a resolution of 1.90 Å.^110 In the α - helical domains, the PDB: 2YCC
structure overlaps almost perfectly with its Fe(II) counterpart (PDB: 1YCC)
(see Figure 7.33 ). Structural adjustments are expressed at the heme and in
movements of internally bound water — water molecules bound near the heme.
For instance, wat166 moves ∼ 1.6 Å closer to the heme ’ s iron ion in the oxidized
species, PDB: 2YCC, Fe(III) – wat166 = 5.0 Å , versus PDB: 1YCC, Fe(II) –
wat166 = 6.6 Å for the reduced protein. Since the heme carries a positive
charge in the Fe(III) oxidation state, wat166 ’ s movement toward the iron ion
may help compensate if its negative oxygen atom dipole points toward the
Fe(III) ion. Other water molecules in the heme ’ s vicinity in the oxidized struc-
ture, wat168 and wat121, form hydrogen - bonding interactions similar to those
seen for the reduced species. In PDB: 2YCC, the side chains of amino acid
residues asn52 and tyr67 form stabilizing hydrogen bonds, with wat166
making this water molecule a major factor in stabilizing both oxidation states
of the heme. Residues 47 – 79, 65 – 72, and 81 – 85, all on the same side of the
protein as iron ion ligand met80, have increased mobility in the oxidized form.
The ligand met80 itself takes slightly different orientations in the reduced
and oxidized forms (see Figure 7.33 ). It thus appears that oxidation - state -
dependent structural changes focus on met80 and the nearby wat166, espe-
cially since wat166 has moved to insert itself within hydrogen - bonding distance
of the tyr67 OH group and met80 ’ s backbone nitrogen. Distances between the
heme ’ s Fe(III) ion and axial ligands his18 and met80 are very similar to those
for the ferrocytochrome (PDB: 1YCC) — that is, N ε 2 – Fe = 2.01 Å and S δ – Fe =
2.43 Å. The heme ligand itself is more distorted from planarity in the oxidized
form (PDB: 2YCC average angular deviation of pyrrole rings from the pyrrole
nitrogens ’ plane = 12.2 ° ) than in the reduced form (PDB: 1YCC, angular
deviation = 9.4 ° ). The normal situation for the heme prosthetic group in these
cytochrome c proteins is to take on a shallow saddle - shape distortion from
planarity. The propionate group attached to the pyrrole A ring, with heme
oxygen atoms O1A and O2A, rotates about the CAA, CBA, and CGB bonds,
resulting in a changed hydrogen - bonding network for the oxidized versus the
reduced proteins. (See Figure 7.32B for the porphyrin atom identifi cation
system.) The differing positions of the heme propionate groups for the Fe(II)
versus the Fe(III) structure are apparent in Figure 7.33. The reference 110
researchers conclude that (1) differences in thermal factors for different aa
residues in the oxidized and reduced forms (as well as increased mobility for
numerous surface residues near axial ligand met80 in the oxidized form) play
a role in promoting electron transfer and in stabilizing the protein in its alter-
nate oxidation states; (2) changing hydrogen - bonding networks induce con-