Molecular chaperones are protein families vital for correct protein folding and preventing aggregation, with various specificities and functions. They are produced in response to stress, particularly heat, and play roles in both folding and degradation processes to maintain cellular homeostasis. Chaperones contribute to disease mechanisms, with disruptions in their function linked to conditions like Alzheimer's disease.

1. CHAPERONES

2. What are Chaperones?

5. The molecular chaperones
are a diverse group of
families of proteins that are
requires for the

6. The chaperones are
concerned primarily
with protein folding
Some chaperones are
non-specific, and
interact with a wide
variety of polypeptide
chains, but others are
restricted to specific
targets.

7. Location
• In humans mostly in Endoplasmic reticulum
• General chaperones: GRP78/BiP, GRP94,
GRP170.
• Lectin chaperones: calnexin and calreticulin
• Non-classical molecular chaperones: HSP47 and
ERp29
• Folding chaperones:
• Protein disulfide isomerase (PDI)
• Peptidyl prolyl cis-trans-isomerase (PPI)
• ERp57

8. The term
`molecular
chaperone`
appeared first in
the literature in
1978
Invented
by Ron
Laskey
History
To describe the
ability of a
nuclear protein
called
‘Nucleoplasmin’
It prevented the
aggregation of folded
histone proteins with
DNA during the
assembly of
nucleosomes

9. • Later extended by R. John Ellis in 1987
o to describe proteins that mediated the post-
translational assembly of protein complexes.
• In 1988, realised that similar proteins mediated this
process in both prokaryotes and eukaryotes.
• Details were determined in 1989, when the ATP-
dependent protein folding was demonstrated in vitro.

10. Properties of chaperones
Molecular
chaperones interact
with unfolded or
partially folded
protein subunits e.g.
nascent chains
emerging from the
ribosome, or
extended chains
being translocated
across subcellular
membranes.
They stabilize non-native
conformation and facilitate
correct folding of protein
subunits.
They do not interact
with native proteins, nor
do they form part of the
final folded structures.

11. • Some chaperones are non-specific, and interact
with a wide variety of polypeptide chains, but
others are restricted to specific targets.
• They often couple ATP binding/hydrolysis to the
folding process.
• Essential for viability, their expression is often
increased by cellular stress.

12. As heat shock proteins
Heat shock
proteins (HSP)
are a family of
proteins that are
produced by
cells in response
to exposure to
stressful
conditions
Several heat
shock proteins
function as
intra-cellular
chaperones for
other proteins
In bacteria like
E. coli,
chaperones are
highly expressed
under high
stress e.g high
temperatures.

13. For this reason, the term "heat shock protein" has
historically been used to name these chaperones.
The prefix "Hsp" designates that the protein is
a heat shock protein.
The reason for this behaviour is that
protein folding is severely affected by
heat and therefore, some chaperones
act to prevent or correct damage
caused by misfolding.

14. Functions of chaperones
• They act as a container for the folding of other protein sub
units as they are called heat-shocked proteins.
• They also prevent the degradation of proteins in spinal stress
conditions.
• Some chaperone systems work as foldases . They support the
folding of proteins in an ATP-dependent manner .(for
example, in the GrpE or the DnaK /DnaJ /GrpE system).

15. • Other chaperones work as holdases . They bind folding
intermediates to prevent their aggregation, for example DnaJ
or Hsp33.
• Chaperones work in coordination by forming assemblies .
• Such assembly chaperones, especially in the nucleus are
concerned with the assembly of folded subunits into oligomeric
structures.
• The first protein to be called a chaperone assists the assembly
of nucleosomes from folded histones and DNA .

16. • They recognize and correct mistakes in folding by
binding to the non polar surface.
• Promote correct folding of their substrate proteins
by unfolding incorrect polypeptide chain
conformations.
• Providing an environment in which correct protein
folding can occur.

17. Macromolecular crowding
• The crowded environment of the cytosol can accelerate the
folding process.
• As a compact folded protein will occupy less volume than an
unfolded protein chain.
• Crowding can reduce the yield of correctly folded protein by
increasing protein aggregation.
• Crowding may also increase the effectiveness of the chaperone
proteins such as Grp E which could counteract this reduction
in folding efficiency.

19. Cell homeostasis
• Two opposite functions
I. Protein folding
II. Degradation.
• The two processes
 Are carried out through the transient formation of
complexes
 Between different chaperones and co-chaperones

20.  Transport across membranes
 Across membranes of the mitochondria
 Endoplasmic reticulum (ER).
 Bacterial translocation—specific chaperone
a. maintains newly synthesized precursor polypeptide
chains in a translocation-competent state
b. and guides them to the translocon.

21. New functions for
chaperones
continue to be
discovered, such
as
a) Assistance in protein degradation
b) Bacterial adhesion activity
c) In responding to diseases linked to protein
aggregation and cancer maintenance.

22. They prevent inappropriate association or aggregation of
exposed hydrophobic surfaces
Direct their substrates into productive
folding
Transport or degradation pathways
Main Role

23. The type I interamolecular chaperones
• First discovered based on the studies on subtilisin, an
alkaline serine protease from bacillus subtilis.
• Mediate the folding of proteins
into their respective tertiary
structures and are mostly produced
as the N-terminal sequence extension.

24. • Mediate the formation of the quaternary or functional
structure of proteins
• Usually located at the
C-terminus of the protein
Type II intramolecular
chaperones

25. Mutations in the intramolecular chaperones can cause misfolding
of the functional domain,
results in distortion of their function leading
to human diseases.

26. Families
• Many families present in eukaryotes and prokaryotes
• Perform many similar and specific functions by
working in coordination systems
• Most common are hsp 70 and hsp 60 families

27. Small heat shock
proteins (hsp25)
[holders]
 Hsp25 is the second
largest of 16
identifiable small heat
shock proteins in the
nematode.
 Protect against
cellular stress
 Prevent aggregation
in the lens (cataract)
Hsp90 ATPase [holder]
 Hsp90 is a specialized
chaperone that assists
in the maturation of
client proteins.
 These proteins
include over a hundred
transcription factors
and kinases, such as
steroid receptors

28. Hsp90
• Hsp90 (HtpG in E. coli) may be the least understood chaperone
• The exact function of Hsp90 is also currently a mystery. Researchers
don't know what it does in the maturation of its client proteins.
• They have discovered that it acts as part of a large complex of different
chaperone proteins.
• Some of these chaperones deliver immature proteins to the complex,
and others assist with folding.
• essential for activating many signalling proteins in the eukaryotic cell
• Each Hsp90 has an ATP-binding domain, a middle domain, and a
dimerization domain

29. Hsp90 (blue) and cochaperone Sba1 (green), with
bound ATP (red).

30. Calnexin , calreticulin
Calnexin (CNX) is a 67kDa integral protein of the
endoplasmic reticulum (ER)
• Calreticulin also known as calregulin in humans is
encoded by the CALR gene.
• Calreticulin is a multifunctional protein that binds
Ca2+ ion rendering it inactive

31. Hsp100 (Clp) ATPase [unfolder]
• The HSP100/Clp proteins are a newly discovered
family, promotion of proteolysis of specific cellular
substrates and regulation of transcription.
• Common ability is to disassemble higher-order protein
structures

32. Hsp100
• Hsp100 (Clp family in E. coli) proteins have been
studied in vivo and in vitro
• Ability to target and unfold tagged and misfolded
proteins.
• Form large hexameric structures
• Unfoldase activity in the presence of ATP.

33.  Proteins in the Hsp100/Clp family form large hexameric
structures with unfoldase activity in the presence of
ATP.
 These proteins are thought to function as chaperones
by processively threading client proteins through a
small 20 Å (2 nm) pore
 Gives each client protein a second chance to fold.
 Forms complexes that are responsible for the targeted
destruction of tagged and misfolded proteins.

34. Hsp 104
• Hsp104 = the Hsp100 of Saccharomyces cerevisiae
• Essential for the propagation of many yeast
prions.
• Deletion of the HSP104 gene results in cells that
are unable to propagate certain prions.

35. Hsp70 chaperones
• Their size is approximately 70,000 daltons
• Best characterized small (~ 70 kDa) chaperone
• Often work in concert with one or more smaller co-
chaperone proteins, which serve to modulate the activity
of the chaperone
• The Hsp70 proteins are aided by Hsp40 proteins (DnaJ in
E. coli), which increase the ATP consumption rate and
activity of the Hsp70s

36. Include DnaK
from the
bacterium
Escherichia coli
The Ssa and
Ssb proteins
from yeast
BiP (for "binding
protein") from the
mammalian
endoplasmic
reticulum

37. • Hsp70 consists of ATP-binding N-terminal domain and
peptide binding C-terminal domain.
• ATP hydrolysis switches off and on the binding ability of C-
terminal domain.
• A special hydrophobic groove formed by α-helices and β-
strands provides the docking site
• For hydrophobic segments of misfolded proteins

38. • Hsp70s crowd around an unfolded substrate, stabilizing it
and preventing aggregation
• Until the unfolded molecule folds properly, at which time the
Hsp70s lose affinity for the molecule and diffuse away
• Hsp70 also acts as a mitochondrial and chloroplastic
molecular chaperone in eukaryotes
• Increased expression of Hsp70 proteins in the cell results in a
decreased tendency toward apoptosis.

40. Hsp 60
also called "chaperonins" are barrel-shaped
structures
Composed of fourteen to sixteen subunits of proteins
that are approximately 60,000 daltons in size
The best characterized large (~ 1 MDa) chaperone
complex

41.  Each subunit has a patch of non-polar amino acid groups
lining the inner surface of the barrel
 This patch recognizes the exposed non-polar amino acids
of misfolded proteins.
 The binding and hydrolysis of ATP triggers
conformational changes within the barrel

43. • Most extensively studied Hsp60 chaperones include
• GroEL and GroES from E. coli
• TRiC/CCT from eukaryotic cells
• TRiC/CCT recognizes a much smaller set of proteins,
and appears to play an additional role in the assembly
of multiprotein complexes

44. GroEL is a
double-ring
14mer with a
hydrophobic
patch at its
opening
GroES is a single-
ring heptamer that
binds to GroEL in
the presence of
ATP or ADP
GroEL and
GroES forms a
well understood
complex

45. GroEL and GroES complex
• GroEL chaperone consists of two rings – cis (or proximal)
upper ring and trans (or distal)lower ring.
• GroES co-chaperone binds to both GroES rings
• Each of the GroEL rings consists of seven identical units
shown in the lower
• The units are arranged in a circular manner and form a
cavity
• GroES upon binding to GroEL serves as “lid”, which
encapsulates the volume inside the cavity

46. Complete their
cycle in 4 phases
Capture (T state)
ATP hydrolysis
(R’’ state)
Encapsulation
Substrate release

48. A top-view of the GroES/GroEL bacterial
chaperone complex model

49. Chaperones andHuman Disease
 It is clear that molecular
chaperones assist with the
folding of newly synthesized
proteins and correct protein
misfolding.
Recent studies now suggest that defects in
molecular chaperone/substrate interactions
may also play a substantial role in human
disease

50.  For example
Mutations linked
to Alzheimer's
disease have
been shown to
disrupt the
expression of
chaperones in
the endoplasmic
reticulum
Several genes linked to
eye degeneration diseases
have recently been
identified as putative
molecular chaperones