UPS Flashcards
define proteostasis
regulation of the level, folding, interactions and localisation of proteins
what are the 3 degradation pathways
cytosolic proteasome (UPS)
lysosomes
ERAD pathway which culminates in UPS
how are the majority of cytosolic proteins degraded
by the UPS pathway
how are proteins targeted to the UPS
ubiquitination
what is ubiquitin
it is a 76aa protein which can be linked to other proteins via it’s Gly76 residue onto lysine residues
describe the process of ubiquitination
e1 formas a thiolesther with the carboxyl group of Gly76 of Ub. E2 transiently carries Ub.
E3 promotes the transfer of Ub onto the substrate protein
which protein in the ubiquitination process is responsible for substrate specificity
E3
describe the structure and function of the proteasome
26S complex formed by a 20S core and 19S cap. it is a compartmentalised protease in which the inner core can hydrolyse bonds. the cap recognises the protein and removes the Ub as well as unfolding the protein to feed it into the core where hydrolysis occurs
what are the 3 proteolytic activities of the 20S proteasome core
tryptic activity cleaves after basic residues
chymotryptic activity cleaves after hydrophobic residues
peptidylglutamylpeptidase activity cleaves after acidic residues
what length are the peptides produced by the proteasome
3-22 aa
how is unfolding of the substrate protein for the proteasome mediated
AAA+ ATPase activity in the 19S cap
what is the role of cytosolic chaperones
recognise misfolded proteins, promote protein refolding, stabalise misfolded proteins
how are chaperones and the UPS linked
cytosolic E3 ubiquitin ligase consitutive Hsc70-interacting protein (CHIP) which binds chaperones Hsc70, Hsp70 and Hsp90 and hence associated with misfiled proteins. CHIP mediated ubiquitination promotes the interaction of CHIP with the 26S proteasome via the co-chaperone BAG-1
what is autophagy
it is the process by which cytoplasmic components are delivered to the lysosome for degradadtion
what are the 3 classes of autophagy
macroautophagy, chaperone-mediated, microautophagy
what are the 3 main enzymes find in lysosomes
cathepsin D cathpsin L and cathepsin B
what is the role of cathepsin B
a cysteine protease whith both endopeptidase and carbocypeptidase activity
what is the role of cathepsin L
a cysteine endopeptidase
what is the role of cathepsin D
an aspartate endopeptidase
what happens to the amino acids produced by proteolysis in the lysosome
they’re transported to the cytosol by membrane transporters
what is the function of macroautophagy
removal of cytoplasmic components e.g. long lived cytosolic proteins, protein aggregates and cellular organelles
what enhances macroautophagy
cell starvation
describe the process of autophagosome formation
cytoplasmic components are surrounded by a double membrane (potentially from ER, golgi, mit or PM). formation requires the action of a number of protein complexes incl. 30AGT proteins in yeast
describe lysosome fusion in macroautophagy
autophagosomes are transported along microtubules to the microtubule organising centre (MTOC). fusion then takes place involving SNARE proteins, VAMP8 and Vti1b
which form of autophagy delivers protein directly to the lysosome
chaperone mediated autophagy
what is the consensus sequence for delivery to the lysosome for chaperone mediated autophagy
KFERQ-like sequences which is normally buried but exposed on misfiled protein
in what conformation can chaperone mediated autophagy degrade proteins
only as single subunits
describe the pathway of chaperone mediate autophagy
Hsc70 recognises signal sequences. proteins are bound by the cytosolic portion of the lysosome membrane protein LAP-2A via the KFERQ motif. this promotes multimerisation of LAMP-2A and then unfolding and translocation of the substrate protein into the lysosome for degradation
what is the hallmark of parkinson disease
levy bodies (mostly made up of a-synuclein) in the cytoplasm of affected neurones
how is aggregation of a-syneuclin spread
it can be transmitted between cells
what is the result of an additional coy of the a synuclein gene
increased risk of parkinsons
how is a-synuclein degraded
both UPS and autophagy although CMA is dominant pathway
what features do neurones of patients with parkinsons disease often have
decreased levels of protein degradation machinery e.g. cathepsin D, LAMP2a and Hsc70 resulting in enhanced transmission of aggregates
how are do the A53T and A30P mutants of a synuclein appear to block the CMA pathway
they contain a KFERQ-like domain which binds Hsc70 for delivery to the lysosome but are not transported into the lumen and impair degradation of other substrates. increased levels of WT protein can have similar effects
why is it thought macroautophagy is also linked to parkinson
in parkinson many cells accumulate autophagosomes demonstrating a failure in clearance
why is mTOR a therapeutic target in parkinsons
activation supresses autophagy. Rapamycin inhibits mTor to promote macropautophay, promoting clearance of a synuclein and reduces neuronal death in animal models
why is TFEB a therapeutic target in parkinsons
it is a transcription factor that regulates macroautophagy and lysosome biogenesis. up regulation promotes clearance of a synuclein and prevents neuronal death
