CYTOCHROMES c 417
In 1990, the Brayer group published the X - ray crystallographic structure of
oxidized (Fe(III)) horse heat cytochrome c at a resolution of 1.90 Å (PDB:
1HRC).^116 The secondary structure of PDB: 1HRC overlaps well with its yeast
iso - 1 counterpart (PDB:2YCC) discussed above. Conformational differences
are noted for (1) residues 22 – 27, part of a surface β - loop, and (2) residues 41 –
43 and 56 – 57, near the heme prosthetic group. The heme groups overlap as
well, both displaying shallow saddle - type distortions. Differences can be seen
in the propionate group attached to heme pyrrole ring A, and this area seems
to exhibit different conformations in most X - ray crystallographic structures
studied. The aromatic side chain group of phe82 is positioned in a co - planar
manner with the heme in both the horse heart and yeast iso - 1 structures. Two
internally located water molecules hydrogen - bond conserved amino acid resi-
dues in slightly different manners for PDB: 1HRC compared to PDB: 2YCC
especially in the area of asn52, the residue that differs most in conformation
between the two structures. Amino acid residues close to wat166 (PDB: 2YCC)
and its counterpart wat112 (PDB: 1HRC) have very similar interactions.
These interactions are collected in Table 7.8 along with axial ligand – iron ion
distances.
In total, fi ve water molecules occupy conserved positions in the structures
of horse heart, yeast iso - 1, tuna, and rice cytochromes c. Three of these are on
the surface of the protein, serving to stabilize local polypeptide chain confor-
mations. The remaining two are internally located. One of these mediates a
charged interaction between the invariant residue arg38 and a nearby heme
propionate. The other is more centrally buried near the heme iron atom and
is hydrogen - bonded to the conserved residues asn52, tyr67, and thr78 (see
Figure 7.33 ). It is shown that this latter water molecule shifts in a consistent
manner upon change in oxidation state if cytochrome c structures from various
sources are compared. The conservation of this structural feature and its close
proximity to the heme iron atom strongly implicate this internal water mole-
cule as having a functional role in cytochrome c ’ s mechanism of action.
Recently, an NMR solution structure of horse heart ferrocytochrome c
(heme Fe(II) ion) in a solvent mix of 70% water – 30% acetonitrile (ACN) was
published (PDB: 1LC1 (minimized average nmr structure) and 1LC2 (30 nmr
structures)).^117 This ferrocytochrome c features a 104 amino acid residue single
chain and an iron - containing heme c prosthetic group. The protein chain has
fi ve α - helices: helix I, residues 3 – 14; helix II, residues 50 – 55; helix III, residues
60 – 69; helix IV, residues 70 – 74; and helix V, residues 88 – 101. The secondary
structure also features two omega loops: turn 1, residues 20 – 35; and turn 2,
residues 35 – 44. The remainder of the chain consists of random coil segments.
The c - type heme is attached covalently to the polypeptide chain through cys14
and cys17. The CXXCH sequence found in cytochromes c displays for PDB:
1LC1 as cys14 – ala15 – gln16 – cys17 – his18. His18 and met80 form the heme ’ s
axial ligands (his18 N ε 2 – Fe = 1.97 Å , met80 S δ – Fe = 2.68 Å ). This is the longest
met80 S δ – Fe distance reported for any structure studied here (see Table 7.8 ).