502
Chaperones, Molecular
other is that the roles of DnaK are so
important that other proteins have evolved
to back them up should DnaK become
nonfunctional. At least one of the roles
for DnaK, namely, that of binding proteins
as they emerge from the ribosome, has
been shown to fall into the latter category.
Another chaperone, called trigger factor,
also has this role, and although loss of
either trigger factor or DnaK alone is not
lethal, a double mutant is not viable under
normal growth conditions (this is referred
to as
synthetic lethality
). In yeast, some
of the Hsp70 homologs are essential and
others are not, and again synthetic lethality
is seen for some combinations of mutants
in different
hsp70
genes, proving that some
of the cellular functions carried out by
Hsp70 are essential. Trigger factor does
not appear to have a homolog in eukaryotic
cells, but another protein called NAC
(nascent-polypeptide association complex)
is thought to fulFll a similar role.
Another important role that has been
demonstrated in
E. coli
and
S. cerevisiae
is not only the prevention of protein
aggregation at heat shock temperatures
but
also
the
breaking
down
of
any
aggregates that may occur. ±or this activity,
another chaperone is required: this is
the protein referred to as ClpB in
E.
coli
and Hsp104 in
S. cerevisiae
.T
h
e
structure and precise mechanism of action
of this chaperone is not known, but it
clearly interacts speciFcally with Hsp70
in mediating protein disaggregation and
subsequent refolding. This interaction is
speciFc: yeast Hsp104 cannot substitute
for
E. coli
ClpB. The role of ClpB will be
considered further below (Sect. 5.2.1).
Hsp40 proteins share many properties
with Hsp70 proteins. In all cases in which
an Hsp70 protein is known to act, an
Hsp40 partner protein is also involved.
4.2.3
Mechanisms of Action of the Hsp70
Family
Both Hsp70 and Hsp40 act by binding to
short, exposed regions of predominantly
hydrophobic stretches in unfolded pro-
teins. Proteins bound to Hsp40 or Hsp70
do not appear to undergo any further fold-
ing, unlike the situation with the Hsp60
proteins, where the protein released from
the complex may be in a very differ-
ent conformation to the one that was
Frst bound by it. Rather, it appears that
the transient binding of these regions is
enough to reduce the risk of their aggre-
gation to other hydrophobic patches on
proteins in the neighborhood. The impor-
tance of this is particularly obvious in those
cases in which proteins have to be pre-
vented from folding until the entire protein
sequence is available to fold, namely, pro-
teins being synthesized on ribosomes, and
proteins being transported across mem-
branes. Similarly, in the case of protein
folding in the ER, proteins that still have
regions exposed and are hence not ready
for transport to the Golgi will bind to the
Hsp70 homolog BiP. This protein has an
ER retention signal at its C-terminus, and
consequently is constantly cycled back to
the ER from the early steps of the secre-
tory pathway, together with any unfolded
proteins that are bound to it.
Although the nature and role of binding
of unfolded protein to the Hsp70 and
Hsp40 proteins are different from those
pertaining to the Hsp60 proteins, the
reaction cycle has some similarities. In
particular, as with the Hsp60 proteins, the
reaction cycle relies upon the chaperones
existing in two states, one with a high
afFnity for unfolded protein and the other
with a low afFnity, with the binding
of nucleotide mediating the transition
between those states. The reaction cycle,
sh
ownin±
i
g
.4f
o
rth
ec
a
s
eo
fth
e
E.
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