CYTOCHROMES c 427
important cytochrome c conserved surface residue phe82 with tryptophan
(F82W, PDB: 2B11), tyrosine (F82Y, PDB: 1B12), isoleucine (F82I, PDB:
2B0Z), and serine (F82S, PDB: 2B10).^133 It is known that mutations at phe82
modestly affect yCc stability, structure, and redox potential^134 and reduce rates
of back electron transfer ( keb ) to the resting state in comparison to ZnCcP
complexed with WT cytochrome c.^135 (See Figure 7.36 and the discussion on
electron transfer in ZnCcP – cytoc complexes that follows.) Substitutions at
phe82 resulted in crystal structures with some interesting differences from wild
type. For instance, F82W ZnCcP – yCc and F82S ZnCcP – yCc mutants maintain
the same contact area between CcP and yCc as wild - type complexes ( ∼ 1208 Å^2
buried surface area), and the F82Y ZnCcP – yCc and F82I ZnCcP – yCc mutants
show CcP to yCc binding at a completely different interface with∼ 803 Å^2 of
buried surface area. Relative to the WT binding, F82Y ZnCcP – yCc and F82I
ZnCcP – yCc mutants show a 90 ° rotation and 8 Å translation of the yCc
domain. Figure 2A of reference 133 illustrates the changes graphically. The
changed domain - binding mode for the F8Y ZnCcP – yCc mutant is surprising
since the difference between phenylalanine (phe,F) and tyrosine (tyr,Y) is
only the addition of the hydroxyl group in tyrosine. One might speculate that
changed hydrogen - bonding scenarios, altered polarity, or local pH variation
could cause the substantial observed structural changes. The F82Y and F82I
mutants show a longer donor (D) to acceptor (A) distance than do wild - type
or F82W and F82S mutants ( ∼ 3 Å greater metal to metal, ∼ 7 Å greater heme
edge to edge, and ∼ 5 Å greater trp191 to heme). The F82W and F82S mutants
show metal - to - metal, heme edge - to heme - edge, and trp191 - to - heme geometry
very similar to those of the WT yeast cytochrome c – ZnCcP complex. The
electron transfer kinetic results are not easy to interpret. First, the back elec-
tron transfer rate ( keb ) is more than 10 times faster than the rate ( ke ) forming
the electron transfer intermediate. Considering only the keb rate, the value for
the yeast wild - type complex vastly exceeds that for any of the mutants (or for
that found for horse heart or tuna wild - type cytochrome c complexes as well).
The fastest keb rates are found for the F82W and F82Y mutants. While the
structure of the F82W ZnCcP – yCc mutant complex is very similar to that
of the WT complex (and therefore electron transfer might be expected to
approach wild - type rates), the F82Y ZnCcP – yCc complex has a substantially
different protein – protein interface (and therefore electron transfer rates might
be expected to be much slower than wild - type). Conversely, the F82S ZnCcP –
yCc mutant, which does have a structure similar to that of WT, exhibits slower
keb rates. The reference 133 authors conclude that the insensitivity of keb to
distance and cofactor (heme) orientation may refl ect limiting dynamic pro-
cesses within the proteins and not just pure electron tunneling.
7.7.4.3 Apoptosis. Apoptosis — programmed cell death — involves elimina-
tion of damaged cells and maintenance of cell homeostasis. Homeostasis can
be defi ned as the ability of living cells to regulate themselves in a dynamic
manner. When apoptosis is deregulated, diseases such as cancers, immune