endoplasmic reticulum

function of ER

• rough ER - synthesis of secreted proteins and membrane proteins

• smooth ER - synthesis of lipids

• folds proteins in the lumen

• glycosylates proteins

• makes disulphide bridges

• oligomerisation

• checks quality of proteins

• calcium store

ER structure

• made up of tubules and flattened sacs - large SA

• linked to outer nuclear membrane

• forms reticulum of interlinked fibres

secretory system

• secretory pathway - pathway of protein trafficking that moves through the ER

• secreted proteins synthesised by ribosomes which dock to ER - mediated by signal sequence at N-terminus

• proteins passed to ER lumen and are modified, then move to Golgi for further modification before being transported out of the cell

secretory pathway - step 1

• docked ribosomes synthesise membrane, lumenal or secretory proteins - ribosome starts reading mRNA in cytosol

• signal peptide synthesised at NH₂ terminus first

• signal peptide recognised by SRP - SRP binds to signal protein to form a complex and docks to SRP receptor in ER membrane

• signal peptide transferred to translocon

• nascent protein synthesised through translocon into ER lumen

signal peptide

• part of newly synthesised protein located at N terminus

• contains one or more positively charged amino acids with an adjacent hydrophobic core

• hydrophobic core prevents aggregation and allows it to insert itself into translocon

signal recognition particle (SRP) and SRP receptor

• SRP:

cystolic ribonucleoprotein hexamer

one subunit can be chemically crosslinked to ER signal peptides to form complex

• SRP receptor made up of an alpha and beta subunit - SRP binds to SRP receptor after forming complex with signal peptide

• P54 SRP subunit and alpha SRP receptor subunit are bound to GTP - they form a pseudosymmetric heterodimer when SRP docks to SRP receptor. This forms 2 complete active sites so GTP can be hydrolysed which destabilises the interface so the heterodimer disassembles

secretory pathway - step 2 for non-membrane proteins

• signal peptides are cleaved by signal peptidases releasing the protein from the translocon into the ER lumen

• translocon - allows passage of polypeptide chain while remaining sealed to small molecules

• signal sequence is docked inside translocon - hydrophobic core forms binding site to allow interactions with translocon

• signal peptidase recognises the amino acid sequence of the signal peptide and cleaves the bond at a specific point - protein released into ER lumen

secretory pathway - step 2 for membrane proteins

• translocon bundles hinge apart to expose hydrophobic binding pocket - hydrophobic core of signal sequence moves in

• protein fed through translocon as it is translated

• protein only goes through upto hydrophobic stop-transfer sequence - signal peptidase recognises hydrophobic stop-transfer sequence and cleaves the amino acid chain

• stop-transfer sequence causes plug peptides to reseal the translocon channel

different ways of protein insertion into membrane

• type - mechanism of insertion, orientation and number of transmembrane domains

• topology of protein determines number of transmembrane domains

• position of positive residues determines orientation

synthesis of type I single pass integral membrane protein

• protein translocated through ER - single pass

• signal sequence cleaved at N-terminus

• stop-transfer anchor sequence stops transfer of the polypeptide through the translocon and becomes the hydrophobic transmembrane segment

• protein moves laterally from the channel to the membrane

• hydrophobic transmembrane segment anchored in interior of membrane

• ribosome is released from translocon once translation is complete

• C-terminus on cystolic side, N-terminus on lumenal side

synthesis of type II transmembrane proteins

• internal sequence preceded by positive amino acid side chains so the N terminus is on the cystolic side

• positively charged AA too charged to enter translocon - translocon hydrophobic

• signal-anchor sequence directs insertion of the polypeptide chain into the ER membrane

• elongation of C-terminal region continues inside translocon - extruded through translocon into ER lumen via cotranslational translocation

synthesis of type III transmembrane proteins

• uses internal signal sequences - no cleavable signal sequence

• positive side chains after signal sequence - C terminus in cytosol

• signal-anchor sequence prevents further extrusion of the chain into the lumen

protein folding in ER lumen

• new proteins for ER-bound ribosomes fed directly into ER lumen

• chaperones mediate folding

• disulphide bonds are added

• oligomerisation occurs

• proteins glycosylated in ER - N-linked glycosylation, makes proteins more hydrophilic to stop aggregation and aid folding, protects proteins from degradation

smooth ER

• connected to rough ER

• exit sites for transport vesicles

• synthesises lipids and steroids

• abundant in cells that metabolise lipids

synthesis of type IV transmembrane proteins

• number of internal hydrophobic transfer sequences determines the number of transmembrane domains

• signal sequence not cleaved