Lecture 16 Endoplasmic Reticulum And Golgi Apparatus Flashcards
Secreted proteins from ER to extracellular space
Synthesised by ribosomes that dock to ER
Mediated by signal sequence at N terminus
Proteins passed into ER lumen
Modified and passed onto Golgi where they are further modified
Finally they are transported out of the cell
Some proteins are synthesed and folded correctly in ER lumen
New proteins from ER bound ribosomes fed into ER lumen.
Chaperones mediate folding (e.g. Nip binds hydrophobic regions preventing aggregation)
disulphide bonds are added by protein disulfide isomerase PDI
Oligomerisation occurs converting reduced protein to oxidised (native) protein. Disulfide bonds stabilise structure in secreted proteins
Step 1: docking to ER
Involves a single peptide (SP) signal recognition particle (SRP)
and SRP receptor
Docked ribosomes synthesise membrane, lumenal or secretory proteins.
Ribosomes start reading mRNA in cytosol
SP is synthesised at NH2 terminus (first)
SP is recognised by SRP
Docks to SRP receptor in ER membrane
Transferred to Translocation channel
(Aka translocon)
Nascent protein synthesised through Translocation channel into ER lumen
Proteins need a channel that is aqueous as they are hydrophilic, cannot pass through membrane directly it’s hydrophobic
Step 2: translocation through ER membrane (soluble excretory proteins)
Signal peptides are cleaved by signal peptidase releasing protein from translocon into ER lumen
Step 2: translocation through ER membrane (membrane proteins)
(type 1) integral membrane proteins start or stop transfer sequences (transmembrane domain) are hydrophobic sequences that remain in the membrane
N terminal sequence is cleaved
Internal sequences become the transmembrane domains
Ribosome synthesises protein until it reaches the transmembrane domain and translocon dissociates
Internal “signal sequence” binds SRP which binds to SRP receptor, targets ribosome to translocon - it is not cleaved
Proteins can be inserted into membranes in several ways
Type 1 - LDL receptor
e.g. influenza HA protein, insulin receptors and growth hormone receptor
Type 2 - Asialoglycoprotein receptor
E.g. transferrin receptor, Golgi galactosyltransferase and Golgi sialytransferase
Type 3 cytochrome P450
Type 4 G protein coupled receptors e.g. glucose transporters, voltage gated Ca2+ channels ABC small molecule pumps, CTFR (Cl-) channels, Sec61
Type 1 single pass integral membrane protein
(see step 2 for membrane protein)
Single pass through membrane
Cleaved signal sequence at N terminus
N terminus on lumenal side
C terminus on cytosolic side
Type 2 membrane protein synthesis
N terminus in cytosol and c terminus in lumen.
No cleaved signal sequence, instead an internal start transfer sequence.
If internal start transfer sequence is preceded by amino acid side chains it’s usually a type 2. This section prevents this part of the protein from entering the translocon and instead a hydrophobic signal sequence is inserted - high charged region excludes N terminus from passing through translocon
Type 3 membrane protein synthesis
Looks like a type 1 but has no cleavable signal sequence instead has a run of charged aminos after it’s transmembrane sequence N terminal lumenal side and C terminal cytosolic side
Type 4 membrane protein
No. Of internal transfer sequences determines no. Of transmembrane domains. Internal sequences bind SRP that binds to SRP receptors which targets ribosome on protein to translocon without cleavage.
Position of pos residues determines orientation of type 4
pos amino sequence pre transmembrane domain - n terminus cytosolic
Pos amino sequence post transmembrane domain n terminus lumenal
Odd no. Of loops leads to each end being on diff sides of the membrane
Summary of soluble secretory protein synthesis in ER
Synthesised by ribosomes
N-terminal signal sequence
Recognised by SRP
sRP recognised by SRP receptor
Docks on translocon
Polypeptide translocated into ER
Signal sequence cleaved by signal peptidase
Folding aided by chaperones
Disulphide bonds formed
May oligomerise
Summary of membrane protein synthesis in ER
Similar to soluble secretory protein synthesis
Some do and some don’t have N-terminal cleavable signal sequence
Have 1 or more internal hydrophobic sequence
Orientation determined/juxtaposed by positive residues
Glycosylation
Proteins are glycosylated in ER - addition of polysaccharides.
Thought to protect proteins from degradation makes them more hydrophilic reducing aggregation and aiding folding
N-linked glycosylation (asparagine, Asn, N)
Starts in ER as protein is synthesised
Smooth ER
Connected to rough ER
Exit site for transport vesicles (maybe?)
Synthesises lipids and steroids
Abundant in cells that metabolise lipids
E.g. Leydig cells in testes- extensive smooth ER to produce testosterone from cholesteron
Function of ER summary
Site for membrane and secretory protein synthesis
Folds proteins in lumen
Glycosylates proteins
Makes disulphide bridges
Oligomerisation
Checks quality of proteins
Calcium store
Smooth er synthesises lipids
Golgi apparatus
Modification of secretory and transmembrane proteins after the ER
completes glycosylation and sorting of proteins so they are packaged in to the right vesicles and sent to the right place
E.g. sent to plasma membrane or endosomes
Discovered by Camillo Golgi
stacks of membrane compartments, flattened sacs with separate internal lumen compartments that each have different enzymes inside as you progress through the stack.
Golgi is orientated all have a cis and trans face:
Incoming cis-face towards nucleus proteins to be edited
Outgoing trans-face towards plasma membrane contains mature proteins
Golgi orientation
Golgi structure
Flat sac-like cisternae
Each is a distinct compartment
Cis face near nucleus incoming site
Trans face near plasma membrane outgoing site
Secretory proteins move in cis to trans direction
Usually near centrosomes in animal cells
Golgi as part of the secretory pathway
Proteins synthesised in ER
Packaged into vesicles
Transported to Golgi
Glycosylated in different stacks
Packaged into vesicles
Transported to plasma membrane
ER vesicle of glycosylated protein fuses to cis Golgi network
Cis Golgi trim oligosaccharides
Medial Golgi attach additions and do further trimming
Trans Golgi add complexity to disaccharides
- enzymes mediate each stage
Retrograde transport Golgi>Golgi and Golgi> ER for retrieval of resident proteins
Vesicle transport between organelles driven by diff vesicle forming “coat” proteins
COPl: Golgi to Golgi and Golgi to ER
COP ll: ER to golgi
Catherine: Golgi to endosome and plasma membrane to endosome
COP ll vesicle formation
Cargo receptors recruit cargo
Sar1 (small gtpase) recruits adaptors to receptor
Adaptors (sec 23/24) bind receptors
Coat proteins (Sec13/31) bend membrane
Golgi summary cisterna progression model
Glycosylates proteins
Series of separate compartments
Recieves new protein from ER in COPll vesicles at cis face
Forms cis network/cisterna by fusion with COP l coated vesicles from cis cisterna
Cis cisterna matures into medial cisterna by fusion with vesicles from medial cisterna
Medial cisterna matures into trans cisterna by fusion with vesicles from trans cisterna
This is the cisternal progression model
