bio230 L10 protein sorting

major functions of the ER

• synthesis and modifications of proteins

• synthesis of lipids

what proteins get sorted to the ER

soluble proteins, transmembrane proteins, proteins destined for golgi, secretion, lysosomes

topological equivalence in terms of vesicular transport

the inside of the ER and the Golgi apparatus are topologically equivalent to the extracellular space

proteins will maintain topological equivalence during vesicular transport — cytosolic segments/proteins in the cytosol, and extracellular segments/proteins in the vesicle

protein sorting to the endoplasmic reticulum

1. mRNA + ribosomes

2. translation starts, ER signal sequence emerges first

3. ribosomes directed to ER membrane

4. co-translational translocation

ER signal sequence

N-terminal, internal, and stop-transfer sequences that direct growing polypeptide chains to the ER

specific hydrophobic sequences, predicted by stretches of hydrophobic amino acids

signal recognition particle (SRP) 

a protein that brings ribosome-amino acid chain complexes to the SRP receptor for protein translocation into the ER

has low affinity for ribosome, hovers outside the exit site until it detects signal sequence

has a high affinity for the ER signal sequence

has a GTPase domain that binds GTP

SRP receptor

a protein that accepts helps the SRP bring the target protein to the translocator

has a GTPase domain that binds GTP

SRP process

ribosome → protein translocator channel (translocon, a gated channel)

ribosome forms a tight seal with the translocator (prevents diffusion of ions, small molecules)

SRP + SRP receptor undergo GTP hydrolysis and complex dissociates

SRP released

protein sorting to the ER: soluble proteins

N-terminal start-transfer sequence binds to the translocator

a signal peptidase cleaves the ER signal sequence

the ER signal sequence laterally diffuses into the lipid bilayer, not seen again. translocator is gated in a 2nd direction

translocated protein is released into the ER

signal peptidase

membrane bound protein that cleaves the ER signal sequence

soluble proteins

a protein that is not associated with the membrane

protein sorting to the ER: transmembrane proteins

single-pass & multi-pass transmembrane (TM) proteins

N and C terminal ends are sorted during translocation

single-pass TM proteins have 3 types of insertions

protein sorting to the ER: transmembrane proteins, singlepass, COOH in cytosol

ER signal sequence: NH2 start-transfer

TM domain is a stop-transfer signal that is cleaved and laterally diffuses into lipid bilayer

protein synthesis continues in the cytosol

protein sorting to the ER: transmembrane proteins, singlepass, positive side in cytosol

TM domain is an internal start-transfer sequence and is not cleaved, laterally diffuses into the lipid bilayer

orientation is determined by the amino acids adjacent to the internal start-transfer sequence

more positive = cytosolic side

membrane protein types

integral (including transmembrane)

lipid anchored

peripheral

formation of glycophosphatidylinositol (GPI)-anchored protein

target protein has c-terminal hydrophobic domain with signal for GPI anchor

GPI anchor is pre-formed in membrane

ER enzyme transfers protein to GPI anchor

GPI-anchored protein ends up on ER luminal side and can go to cell exterior surface