36
Aggregation, Protein
allowing the release of DnaJ and P
i
.In
the following step, GrpE acts as an ex-
change factor to regenerate the ATP-bound
state of DnaK. The unfolded polypeptide
chain is released into the bulk solution.
Thus, Hsp70 systems bind and release the
polypeptide in an unfolded conformation.
The unfolded protein has the possibil-
ity either to fold or to be transferred to
the GroEL system, as illustrated in Fig. 6.
Signi±cant insights into this mechanism
were obtained from structural data. The
three-dimensional structures
of
Hsp70
andDna
Ja
swe
l
la
stho
seo
facomp
lex
(a)
(b)
Fig. 7
Crystal structure of
GroEL–GroES–(ADP)7 complex
determined by Sigler et al. (a) view
along the axis and (b) view from the top
of the complex. (Reproduced from the
PDB web site.)
DnaK–polypeptide and a complex of GrpE
with the ATP binding domain of DnaK
are known. DnaK and its homologs are
composed of two domains, a C-terminal
domain that binds ATP and an N-terminal
domain that binds peptides. GrpE is a
tight homodimer associated along two long
helices. It binds DnaK–ATPase domain
through its proximal monomer. DnaJ ac-
tivates the ATP hydrolysis by DnaK. It
was shown that a conformational change
may occur upon ATP binding, opening the
polypeptide binding cleft in the polypep-
tide binding domain of DnaK. The closed
state may correspond to the ADP-bound
conformation. The ADP-bound state of
DnaK binds the peptide tightly. Peptide
release requires the dissociation of ADP,
which is mediated by GrpE. DnaK then
rebinds ATP.
The GroEL–GroES system acts by a dif-
ferent mechanism in which the unfolded
protein is sequestered. The chaperonins
are large cylindrical protein complexes.
The crystal structure of
E. coli
chaperonin
GroEL was determined in 1994 and that
of the asymmetric GroEL–GroES–(ADP)7
complex in 1997 by Sigler and his group.
GroEL consists of two heptameric rings
of 58-kDa subunits stacked back to back
with a dyad symmetry and forming a
porous cylinder (Fig. 7). Each subunit is
organized in three structural domains. A
large equatorial domain forms the founda-
tion of the assembly and holds the rings
together. It contains the nucleotide bind-
ing site. A large apical domain forms
th
eendo
fth
ec
y
l
ind
e
r
.Th
eap
i
c
a
ld
o
-
main contains a number of hydrophobic
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