ER protein folding and degradation Flashcards
where is the signal sequence for proteins located in the ER
N-term
which protein recognises signal sequences on proteins targeted to the ER
cytosolic ribonucleoprotein complex signal recognition particle (SRP)
describe the role of SRP and the process of translocation of nascent proteins into the Er
it binds signal sequences and halts translation to prevent folding in the cytosol. it delivers the ribosome and nascent chain to the ER membrane via interactions with the SRPR. SRP is displaced once the ribosome binds the Sec61 translocon and translation resumes. sec61 forms an aqueous pore to translocate the protein. folding and modification occurs as the protein is translocated
what is BiP
it is an ER luminal Hsp70 required for translocation, folding and degradation
how is BiP involved in protein translocation into the ER
it binds the nascent chain as it is inserted and retains it in the ER
how is BiP involved in protein assembly in the ER
energy from ATP hydrolysis enables BiPto bind unfolded regions of proteins. exchange of ATP for ADP causes dissociation and provides an opportuntiy for the protein to fold
why does disulphide bond formation occur in the ER
it is an oxidising environment and favours formation
at what point can disulphide bond formation occur in the ER on nascent proteins
it can begin when the protein is being translocated
how is disulphide bond formation promoted between correct pairs of cysteine
protein disulphide isomerases can oxidise, reduce and rearrange disulphide bonds
what occurs in N-linked glycosylation
a preformed oligosacharride is transferred from a dolichol lipid precursor to asparagine (N) by oligo-sacharyltransferase (OST)
what is the preformed oligosaccharide chain formed of which is added in N linked glycosylation
2 N-acetylglucosamines, 9 mannose, 3 terminal glucose
what is the consensus sequence for N linked glycosylation
Asn-X-Thr/Serwhere x=any amino acid except proline
what are the roles of N-linked glycosylation
improves protein solubility, provides binding sites for challenging and calreticulum which facilitates interactions with PDIs and finally it is used to monitor protein folding
after addition of the N-linkedoligosacharride what happens to it
it is trimmed to leave a single glucose residue by sequential action of glucosidases I and II. the trimmed oligosacharride is a ligand for calnexin and calreticulum which retain the protein in the ER to prevent aggregation and promote folding by binding PIDs. the final glucose is then trimmed which causes the protein to dissociate from calnexin and calreticulum.
what happens to a protein if, after N-linked glycosylation and consequent trimming and folding, the protein is still not correctly folded
a glucose residue is added by UDP-glucose glycoprotein glycosyl-transferase (UGT) and the cycle is repeated. alternatively a slow process of mannose removal catalysed by mannosidases can occur which reduces action of UGT. if sufficient mannose residues are removed the protein is diverted from the folding pathway to the ERAD pathway
what proteins recognise misfolded proteins in the ER
adaptor proteins- SEL1L which can also recruit additional adaptors incl. lectins XTP3-B and OS9. also BiP
where does ubiqitination of proteins destined for the ERAD pathway occur
the cytosolic face of the ER membrane
name an example of a ER membrane bound E3 ubiquitin ligase
HRD1
why is SEC61 a candidate for the retro-translocation pore in the ERAD pathway
because mutations can impact on getting proteins out of the ER
why is E3 HRD1 a candidate for the retro-translocation pore in the ERAD pathway
it forms multimers in the ER membrane and a simplified system showed that this was sufficient for retrotranslocation
what are derlins
membrane bound proteins which promote luminal protein transport across the membrane
after ubiquitination how is the protein retrotranslocated
it is recognised by the cytosolic AAA+ ATPase p97/Cdc48 which uses ATP hydrolysis to energise the extraction of the protein from the ER into the cytosol
how are proteins in the ERAD pathway degraded
they are delivered to the proteasome in the cytosol
how might translocation and degradation in the ERAD pathway be coupled
because there is a population of proteasome at the cytosolic face of the ER membrane and they may act to pull the protein from the membrane via the AAA+ ATPase activity in the 19s cap
what triggers the UPR
accumulation of misfolded proteins in the ER
what effect do the signals induced by the UPR have overall
they reduce bulk protein synthesis, promote chaperone production, increase ERAD and increase the amount of ER
which proteins are unregulated as a result of the cleavage of AFT6 to form the AFT6(N) transcription factor in the UPR
BiP, GRP94 (an Hsp90), PDI
what cellular pathways does XBP1 affect as a result of cleavage by IRE1 activation in the UPR
it promotes regulation of lipid biosynthetic enzymes, up regulates ERAD components, induces plasma cell differentiation and degrades mRNAs of secretory proteins thus reducing protein synthesis and producing more ER
what is CFTR
an ATP gated plasma membrane channel, expressed in epithelial cells, which transports Cl- ions across the membrane into the mucous to increase water movement and make it less viscous
what occurs in the absence of CFTR function in the lungs
there is a build up of viscous mucous resulting in inflammation and infection causing lung damage
what is the most common mutation that occurs in CFTR
deletion of F508 (known as DF508)
what is the fate of the DF508 mutant
despite retaining some functionality, 100% of it is retained in the ER and regarded via the ERAD pathway
describe UPR dependent trafficking of CFTR DF508 to the cell surface
under ER stress, unconventiona protein secretion dependent on GRASP is activated. IRE1 signalling activates GRASP which binds to CFTR DF508 and targets it to the PM.
although UPR dependent trafficking of CFTR DF508 to the PM may be used as a therapeutic, what are some of the problems faced by this potential solution
there are QC mechanisms at the PM which may recognise the misfolded protein and recruit chip leading to degradation
aside from utilising the UPR dependent trafficking system, what other potential therapies are being researched for CF
small molecules which can stabilise the misfolded protein to increase expression at the cell surface. this is not sufficient by itself but in conjunctio with another drug to cause channel opening may increase function
what are the HCMV US2 and US11 proteins
they are ER localised integral membrane proteins which target MHCI for degradation by acting as virally encoded adaptors for the ERAD pathway
what is the effect of choler toxin on the body
it promotes electrolyte and water movement into the intestinal lumen resulting in severe diarrhoea
what is the molecular mechanism of cholera toxin
it is internalised by cells and trafficked to the ER via the secretory pathway. Here the CTA1 subunit dissociates from CTA2/CTB dependent on PDI. CTA1 is unfolded and retrotranslocated into the cytosol. it escapes proteasomal degradation due to a lack of sine residues for ubiquitination. in the cytosolic binds adenylate cyclase increasing levels of cAMP which activates PKA. PKA phosphorylates CFTR promoting Cl- secretion
