416 IRON-CONTAINING PROTEINS AND ENZYMES
transfer proteins will exhibit variations of several orders of magnitude ink 11 ,
depending on their changed electron transfer (eT) reactivity. The eT reactivity
can then be related to structural changes (as found in X - ray crystallographic
studies) and thus to eT mechanisms for WT versus mutant cytochromes c. In
other words, variations in eT reactivity can be interpreted in two ways: (1) in
terms of the effects of mutation on native oxidized or reduced cytochromes c
(electrochemical effects) or (2) in terms of the mechanism by which proteins
change oxidation state (kinetic effects). The results for the surface variants
phe82X, where X = tyr, leu, ile, ala, ser, gly, indicated faster eT rates as X
decreased in size. All variants exhibited faster rates than did wild - type. Faster
relative rates were found for reduction of ferricytochrome c by [Fe(EDTA)] 2 −
than for oxidation of ferrocytochrome c by [Co(phen) 3 ] 3+. The reference 114
authors interpreted the size relationship as favoring faster eT when a closer
approach to the redox partner was possible. The researchers also discovered
that, at least by this kinetic method, changes in the internal hydrogen - bonding
network did not translate to lowered electron transfer reactivity. In fact,
for the Y67F mutant discussed above, the relative rate of electron transfer
(k 11 corr) was nearly 11 times faster than that for wild - type ferri - or ferrocyto-
chrome c.
The extended study of yeast iso - 1 - cytochrome c by these researchers leads
to several conclusions about the protein: (1) In all studied mutants, the overall
cytochrome c fold is unchanged; (2) his18 and met80 are retained as the iron
axial ligands in all studied mutants; (3) in the native (wild - type) protein, move-
ment of the wat166 molecule closer to the heme iron in the Fe(III) state and
its reorientation to bring its negative dipole closer to the heme iron ion will
help relieve positive charge on the heme prothestic group (with two negative
charges on the propionate group the Fe(II) heme is uncharged); (4) breaking
of the tyr67 OH – met80 S δ hydrogen bond in the Fe(III) state allows the sulfur
lone - pair electrons to help balance the heme group ’ s +1 charge; and (5)
changes in hydrogen - bonding patterns near the heme are focused near con-
served (invariant over different biological species) residues and the heme
pyrrole A ring ’ s propionate group.
7.7.3 Mitochondrial Cytochrome c (Horse),
One of the fi rst X - ray crystallographic studies of horse heart cytochrome c was
published by Takano and Dickerson in 1980.^115 (PDB: 3CYT) These workers
fi rst noted the small but signifi cant ferro - and ferricytochromes c conforma-
tional differences surrounding the buried water molecule that is hydrogen -
bonded to aa residues asn52, tyr67, and thr78 (see Figure 7.33 ). They saw, in
the oxidized state, that the water molecule moves closer to the heme while the
heme moves slightly out of its heme crevice, leading to a more polar heme
environment. They noted that the cytochrome c heme crevice is bounded by
lysines 8, 13, 27, 72, 79, 86, and 87.