420 IRON-CONTAINING PROTEINS AND ENZYMES
The heme iron atom is 0.16 Å out of the plane formed by the four porphyrin
nitrogen ligands and bulges toward the met80 sulfur ligand. Heme pyrrole ring
C, which has been proposed to function as part of the physiological electron
transfer pathway, exhibits pronounced buckling in the PDB: 1LC1 structure.
It is the closest pyrrole ring to the surface residue phe82, also implicated in
electron transfer pathways. The reference 117 authors compare their structure
in 30% nonaqueous solvent with a horse heart ferrocytochrome (Fe(II)) solu-
tion structure in aqueous solution published by the I. Bertini group in 1999
(PDB: 1GIW (minimized average structure) and 2GIW (40 NMR struc-
tures)).^118 In Figure 7.34 , the PDB: 1GIW structure is visualized in green, while
the PDB: 1LC1 structure is visualized in colors: helix I, yellow; helix II, red;
helix III, magenta; helix IV, cyan; and helix V, royal blue. Turn 1 for PDB: lLC1
is colored orange, while turn 2 is colored gold. Random coil segments for both
structures are visualized in green.
The α - helical portions of the two molecules overlap almost completely,
while the loop portions show considerably more variation. Using NMR ana-
lytical methods, the reference 117 (PDB: 1LC1, 1LC2) authors indicate higher
RMS (root mean square) deviations, and thus higher fl exibility, for the back-
bone atoms in segments 21 – 24 and 41 – 45 for the ferrocytochrome in 30%
ACN than for the ferrocytochrome in aqueous solution. (Compare the orange
and gold loop sections for PDB: 1LC1 to the PDB: 1GIW counterparts in
green in Figure 7.34 .) Polar amino acid residues on the two ferrocytochromes
c surfaces show considerable conformational change, although their backbone
atoms remain in similar positions. In Figure 7.34 , these residues (lysines 8, 60,
72, 86, 87, leu32, and his33) are labeled. In the ferrocytochrome in 30% ACN
solution, the polar residues are more bent in toward the protein probably
because of the unfavorable reaction of the polar side chains with the less polar
(than water) solvent medium. The aromatic ring orientation of the important
surface residue phe82 is different in the two structures, with the PDB: 1LC1
residue ’ s ring being situated more in parallel with the heme plane. A notable
change in the conformation of his33 confi rms other studies showing that met80
is removed as a ligand in cytochrome c denaturing solutions and that his33
can become an iron ion ligand in these circumstances.^119 These authors have
proposed his33 to be the sixth (and second axial) ligand in the V form of
ferricytochrome c in 30% ACN solution and an alternative axial ligand replac-
ing met80 in cytochrome c denatured by urea.^120 Similarly, lys72 exhibits a
conformational change that may allow it to become a sixth ligand in the IVa
form of of alkaline ferricytochrome c and the IVa form of ferricytochrome c
in mixed solvents. The nomenclature for conformationally changed ferricyto-
chromes c includes: (1) state III — the native conformation that prevails at
neutral pH; (2) state IV — conversion from state III at p Ka values between 8.5
and 9.5; and (3) state V — forms at alkaline pH values greater than 9.5.
The conformational changes have been studied by a number of different
methods, including resonance Raman (RR) spectroscopy by Mauk and
co - workers.^121