ER Protein Targeting: L12

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Questions and Answers

What is the core characteristic of ER-localised proteins?

A mixture of polar and non-polar amino acids throughout the sequence

An absence of hydrophobic amino acids

A sequence of basic amino acids followed by hydrophobic amino acids

A core of ~8 hydrophobic amino acids preceded by several basic amino acids (correct)

What role does the Signal Recognition Particle (SRP) play in protein localization to the ER?

It assembles into a complex with ribosomes to synthesize proteins

It binds to the signal sequence through a hydrophobic pocket (correct)

It transports proteins directly into the ER lumen

It cleaves the signal sequence from proteins after they reach the ER

Which amino acid property is typically found at the N-terminal signal sequence of proteins that localize to the ER?

Hydrophobic core preceded by basic amino acids (correct)

High hydrophobicity with a mix of polar amino acids

Hydrophobic core followed by acidic amino acids

High polarity with charged side chains

In the context of preproinsulin localization, what happens to the signal sequence after it directs the protein to the ER?

It is cleaved between specific residues Signup and view all the answers

Which of the following proteins is NOT mentioned as having an ER localization signal?

Mouse cytochrome C Signup and view all the answers

What characterizes the process of protein import to the Endoplasmic Reticulum (ER)?

It is a co-translational process involving membrane-bound ribosomes. Signup and view all the answers

Who received a Nobel Prize for the discovery related to intrinsic signals in proteins?

Blobel and Sabatini Signup and view all the answers

What happens to proteins during their import into the ER?

They are shortened as they are imported. Signup and view all the answers

Which component forms a continuous sheet that encloses the ER lumen?

Rough ER and smooth ER Signup and view all the answers

What is a significant feature of the rough ER?

It has membrane-bound ribosomes attached to its surface. Signup and view all the answers

What does the Signal Hypothesis explain regarding proteins?

Proteins have intrinsic signals allowing them to reach their destinations. Signup and view all the answers

Which cellular structure is involved in the mechanism of ER translocation?

Ribosomes Signup and view all the answers

What is one crucial function of the Endoplasmic Reticulum?

Synthesis of protein and lipids Signup and view all the answers

What is the primary role of the SRP receptor on the ER membrane?

To temporarily block protein translation Signup and view all the answers

Which process uses GTP as energy for conformational changes in the SRP?

Changing the conformation of the SRP Signup and view all the answers

What mechanism does the Sec61 translocon use to translocate proteins across the ER membrane?

Gated channel mechanism Signup and view all the answers

What is the function of the hydrophobic stop transfer sequence during protein translocation?

Terminate translocation and retain proteins in the ER membrane Signup and view all the answers

How does the RUSH assay assist in measuring ER trafficking?

Using selective retention mechanisms Signup and view all the answers

What is the role of chaperones like BiP in protein translocation?

They assist with protein folding and translocation Signup and view all the answers

Which component is responsible for the recognition of the ER signal sequence during translocation?

SRP and Sec61 Signup and view all the answers

Which type of proteins are predominantly synthesized from the outer layer of the ER membrane?

Class 1 LD proteins Signup and view all the answers

What feature distinguishes amphipathic helices?

They have a specific distribution of charged and hydrophobic residues Signup and view all the answers

During protein translocation, which part of the protein is recognized first by the SRP?

N-terminal hydrophobic signal peptide Signup and view all the answers

Which sequence is crucial for retaining proteins in the ER?

C-terminal retrieval sequence Signup and view all the answers

What is the function of the Sec61 translocon during protein translocation?

To provide a channel for protein movement across membranes Signup and view all the answers

Which mechanism allows for the release of fully synthesized proteins into the ER membrane?

Opening of the lateral pore of the Sec61 translocon Signup and view all the answers

What energy source is utilized during the cycles of chaperone binding for protein translocation?

ATP Signup and view all the answers

Study Notes

BIOL2056: Cell Biology - Protein Targeting: ER

Recommended Literature: Alberts Molecular Biology of the Cell, 6th edition, Chapter 12
Lecturer: Dr Triana Amen, [email protected], Building 85, Room 3035
Learning Objectives:
- History of protein targeting discovery (Signal Hypothesis)
- Role of protein synthesis in protein import to the ER
- Mechanism of ER translocation
- Protein import into Lipid Droplets (LDS)
- Case study: measuring ER trafficking

Endoplasmic Reticulum (ER)

ER and nuclear membranes form a continuous sheet enclosing a single internal space, the ER lumen
ER is composed of rough ER (studded with ribosomes) and smooth ER
The ER lumen is the space inside the ER
ER membranes and the nuclear membranes are continuous

ER Protein Import

A co-translational process: protein synthesis and import into the ER happen simultaneously
Membrane-bound ribosomes coat the membranes of rough ER
Proteins have a signal sequence (hydrophobic amino acids) at their N-terminus that targets them to the ER
Signal peptidase cleaves the signal sequence within the ER
This signal sequence is recognized by the signal recognition particle (SRP)

Signal Recognition Particle (SRP) and SRP Receptor

Initially, SRP binds to the signal sequence on the nascent polypeptide chain as it emerges from the ribosome
SRP binding halts protein synthesis
SRP-ribosome complex moves to the ER membrane
The SRP receptor on the ER membrane recognizes SRP
SRP detaches, and the ribosome binds to the Sec61 translocon
Protein synthesis continues, and the polypeptide chain enters the ER lumen
GTP hydrolysis drives the SRP cycle

Sec61 Translocon

A protein-conducting channel in the ER membrane
Gated by a short alpha helix, keeping the membrane impermeable to small molecules
The channel opens transiently to translocate the polypeptide chain
The pore can also open sideways, important for transmembrane proteins and signal peptide release

Translocation into the ER Lumen

Requires energy provided by GTP hydrolysis during SRP binding and conformational changes
ATP-dependent chaperones (like BiP, Hsp70) help with translocation and protein folding.
Proteins synthesized in the cytosol and destined for the ER lumen have an N-terminal signal sequence targeting the ER, aided by SRP, which halts translation until it reaches the ER.
Additional hydrophobic Stop-transfer sequence to stop translocation into the lumen, embedding proteins in the ER membrane

Polyribosomes for Efficient Translation

Multiple ribosomes can bind to a single mRNA molecule, forming a polyribosome
This ensures efficient synthesis of multiple copies of the same protein
The mRNA encoding a protein targeted to ER remains membrane-bound

Protein Localization to Lipid Droplets (LDS)

LDS are produced from the outer layer of the ER membrane
Two classes of proteins associate with LDS:
- Class 1 proteins, containing hydrophobic hairpins
- Class 2 proteins containing amphipathic helices or lipid anchors
Class 1 proteins enter the ER and are directed to the LDS.
Class 2 proteins often are inserted post-translationally after being imported into the LDS

Retrieval Sequence (KDEL)

Some ER proteins contain a retrieval sequence (KDEL)
This sequence binds to receptors in the ER, preventing the proteins from being transported to other organelles.

Case Study (RUSH Assay)

A method for measuring ER trafficking.
Different components are fluorescently tagged, and changes in localization over time are monitored using fluorescent microscopy

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