Download Protein Folding and Quality Control

Lecture 24
Protein Folding and Quality Control
Folding
Function: making specific functional domains critical for function (occurs following or coincident
with synthesis)
Sequence dependence: Final structure of protein is
dependent on amino acid sequence and properties of amino
acids that make up polypeptide being synthesized. Proteins
will fold during synthesis to achieve lowest possible energy
state. Hydrophobic amino acids will group together forming
hydrophobic interactions and a hydrophobic core while
hydrophilic amino acids will go outside, interact with water
and other water soluble molecules increasing solubility of the amino acid.
Refolding: Following denaturation, many proteins can refold when put back into ideal conditions.
HOWEVER, there are many proteins that cannot reform and need help along the way.
Heat shock proteins: they are molecular chaperones.
Problem: at high temperatures, hydrophobic amino acids
do not interact properly, don’t form core, interact with other
proteins and form aggregates. Solution: HSP aids in regular
folding of protein. HSP increase rapidly in presence of heat or
other cellular stresses. Method: 1) When in ATP bound state
binds to nascent unfolded polypeptide 2)ATP hydrolyzed and
HSP form hydrophobic pockets, allows for normal folding of
hydrophobic elements. 3+4) When ATP reassociates, goes
back to original configuration and folded protein is released.
GroEL : acts as bacterial chaperonin (large
macromolecular machine devoted to folding) like TriC in
humans. Structure: combination of 2 rings, repeated
subunits. Barrel structure with interior. 2 different states.
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Function: 1)In tight conformation, can accept
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polypeptides. Allows polypeptide’s hydrophobic residues
to interact. Leads to release of properly formed
polypeptide. 2) Relaxed conformation allows polypeptide
to leave the barrel and do its process. Relaxed by
association of GroES and ATP.
Disulfide bridges: Function: facilitate appropriate folding and
require oxidation of SH groups on cysteines. ER dependence:
proteins for processing only present in ER (endoplasmic reticulum) so
proteins with S-S bridges must go through ER. Which cells: Proteins
to ER are for excretion or membrane. Cells that excrete a lot of
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protein have a lot of PDI (ex. Pancreas that secretes insulin) Method:
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1) PDI recognizes thiol groups. 2) PDI has reduced active site, so
catalyses reaction that oxidizes (removes Hydrogen) from cysteine
forming bridge this involves: a) electron transfer from thiol group b)
oxidized thiol acting on neighbour forming bridge 3) PDI rearranges
bridges if necessary (if protein is not in optimal state after sequential
formation of bridge) 4) Ero 1 brings PDI back to oxidized state
so that it can be used again (since it was reduced by rxn). IMPORTANT: all proteins with
disulfide bridges contain cysteine but NOT ALL CYSTEINE CONTAINING PROTEINS FORM
BRIDGES. They MUST be brought to ER to form a bridge.
Plaques: Incorrect folding can lead to pathologies involving aggregates of proteins
forming plaques: Alzheimers & Huntington’s disease. Alzheimers: Protein
aggregates or Amyloid plaques (misfolded amyloid proteins) and tangles present in
the diseased brain. Huntington’s: Protein aggregates or plaques arise in neurons
within the brain. These are associated with neuronal cell death (necrosis).
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Molecular Biology
Lecture 24
Kuru: terrible disease afflicting Fore people. Members of the tribe,
particularly women and children, ate parts of the relative. Neuronal
tissue was one of first to be consumed. Pathogen was a small protein
(prion). Associated with laughing death. People were having ongoing
neuronal degradation, and eventually death. Loss of neuronal function.
Stopped cannibalism and got rid of disease. Prion disease: related to
scrapies, BSE (mad cow disease), Creutzfeldt-Jakob
Prion protein (PrP): 2 states: PrPc
(conformer state) is non-infectious.
Structure: 3 alpha helices. PrPsc (non
conformer state) is infectious. Structure: 2
alpha helices + 1 beta sheet. DIFFERENT
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STRUCTURE DIFFERENT FUNCTION.
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Amino acid sequences of both forms are
identical. Infection: PrPsc is highly protease resistant thus
accumulates and form plaques and results in neuronal cell death.
Also, PrPxc can convert PrPc to infectious form! PrPsc thus acts in a
dominant manner and causes infection.
Quality Control
Maturation, export, and pioneering round: involved in
translational quality control. a) SR proteins: define exons for proper
excision of introns. b) Poly Adenylation. c) Export actors: loaded
onto mRNA to bring out of nucleus. All loaded factors removed or
you get NMD!!!!
NMD (nonsense mediated decay): Wild type situation: proteins
knocked off sequentially. Mutant type situation: In frame stop
within exon. Ribosome drops off mRNA but still has proteins
associated with it. Ex. if had something with only ligand binding
domain, could be a dominant negative protein (completes half of
function so blocks normal protein from completing function). By
keeping SR and other proteins associated to it, tells cell is a bad situation. NMD machinery is
set up to chew up mRNA because don’t want to translate that protein.
Endoplasmic Reticulum
Earmarking: proteins are earmarked for numerous destinations. Info for
localization found within polypeptide sequence. Cytosol: contain little
localization info so stay in cytoplasm. Nucleus: have a nuclear localization
sequence. Mitochondria, chloroplast, peroxisome: other signalling
information. ER: secreted proteins come here by recognition of specific
sequences called a signal recognition sequence.
Rough ER: ribosomes are bound to ER (same as
free ribosomes) and introduce growing polypeptides into ER.
SRP: recognizes signal sequence and 1) brings
down polypeptide 2) inserts it into ER
3) polypeptide modified by proteins in ER (step 3
not related to SRP per se)
Ire1 dimerization: important for ER unfolded
protein response (UPR) Proteins involved in this
are BIP and IRE (ER chaperones that ensure
proper folding thus preventing aggregation and
binding irreversibly to misfolded proteins). IRE
normally in monomeric situation associated with
BIP. BIP is a molecular chaperone within the ER.
BIP thought to bind to IRE so can’t form dimer
when too much protein unfolded. When sequestered away from IRE, can
dimerize and increases transcription.
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Molecular Biology
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Alternative explanation: Unfolded protiens forms scaffold
that allow to form dimers, through inherent ability to snip
transcription factor mRNA, and increases transcription of
things that are for folding mrna. Based on structural
biology. Perhaps this case with BIP is not the best
explanation. IRE is binding to proteins themsleves and
that is forming the dimerization.
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Molecular Biology