Lecture 6: Intracellular Compartments and Protein Sorting
- Many proteins need to enter ER for modification w/ sugars. This occurs at same time they are being synthesized
by ribosomes. Translation begins w/ synthesis of short signal peptide sequence, a signal recognition particle,
protein complex binds to peptide which translation continues. SRP then binds to receptor in ER membrane
anchoring ribosome which binds its receptor and the signal peptide meets the translocator. Translation
continues and protein passes through translocator. The signal peptidase cleaves the signal peptide leaving the
new protein molecule in lumen of ER.
- Signal sequences are a stretch of AA seq of protein that directs protein to correct location in cell. In gene,
located at beginning – 5’ end. Found at N terminus of protein.
- A signal peptidase can remove signal from protein
- Signal sequences can be internal stretches of AA which remain part of protein
- Sorting of a secreted protein:
o Translation starts on cytosolic ribosomes
o Signal sequence at amino terminal end directs protein to ER
o The signal sequence is hydrophobic – it interacts w/ translocator. After it is cleaved, it ends up in
membrane of ER which is then degraded by other proteins. This is known as cotranslational
translocation to ER b/c it happens while the protein is still being translated.
o Protein is inserted through membrane by a protein translocator
o Signal sequence cleaved, left being in ER membrane, secreted protein ends up in ER lumen
o Secreted protein moves in transport vesicles via secretory pathway
o Released by exocytosis at the plasma membrane. The vesicle fuses with plasma membrane (secretion)
- Transmembrane proteins do not get secreted; you need it to stay part of membrane. Once its been synthesized
it stays in membrane and it is transferred via vesicles. Some proteins will eventually end up as membrane
spanning protein (Transmembrane protein). Start transfer sequence: targets protein to ER which pass through
translocator and it binds/ interacts w/ translocator protein and protein continues to be translated.
- Stop transfer sequence: hydrophobic. Binds hydrophobic that stretch of 20-30 AA which tells translocator is
- Other parts are of protein are hydrophilic, translocator opens in diff manner – can open sideways and release
content in membrane (mature protein and start sequence)
- There are 2 diff ways a single pass Transmembrane protein can be oriented in ER membrane.
o For a protein with an internal signal sequence.
o Hydrophobic. Interacts with translocator – can interact in 2 diff orientations depending on seq od AA
around signal seq
o Eg.1: bind to translocator such that amino side stays in cytosol. COOH in ER lumen
o Eg. 2: bind to translocator such that AA side in ER lumen, COOH stays in cytosol
o What determines direction?
AA around seq signals are + or – charges. Depending on where the more –ively charged AA lies
– direct how protein will interact with translocator. If more –ive AA are after signal seq. on
COOH end of seq – that will direct protein to be inserted so COOH directed toward ER lumen.
- A multipass Transmembrane protein: (how they get inserted into membrane): has 2 membrane spanning
domains. Have 2 stretches of hydrophobic AA – one is start transfer seq which directs protein to ER and binds
translocator and starts transfer of seq across membrane. Another stretch of AA is translated into protein and
that acts as a stop transfer sequence which anchors both start and stop sequences in translocator which is then
able to open laterally and release protein into membrane into Transmembrane spanning domains.
o In this process, the start transfer sequence is more active b/c it directs the protein to the ER. Stop
transfer sequence is passive b/c it has hydrophobic stretch on AA that translocator recognizes b/c it is
being pulled through translocator.
- Path of Transmembrane protein from translation to PM:
1. Cytosol to ER to Golgi by a vesicle that buds off and fuse with golgi and transfer protein to its
o In golgi the proteins get their complex glycosylations and glycosylation of lipids also occurs.
2. Golgi PM. Vesicles fuse and transfer proteins into plasma membrane
- Golgi apparatus: receives proteins and lipids from endoplasmic reticulum, modifies them, and then dispatches
them to other destinations in the cell. It has stacks of cis cisterna. The move in organized manner. They start in
ER, cis with golgi network (CGN) and then move through all of individual golgi cis cistern by budding and fusing.
Then at trans golgi (trans face) they bud off again.
o Process of glysocylating is complex. It requires many enzymes that add on and modify sugars as proteins
mature. This is organized in golgi