Protein Sorting: Targeting of Proteins to Organelles

Protein Sorting: Targeting of Proteins to Organelles

Protein sorting ,

Introduction of protein sorting

Protein targeting or protein sorting is the mechanism by which a cell transports proteins to the appropriate positions in the cell or outside of it.
Sorting targets can be the inner space of an organelle, any of several interior membranes, the cell’s outer membrane, or its exterior via secretion.
This delivery process is carried out based on information contained in the protein itself.
Correct sorting is crucial for the cell; errors can lead to diseases.
Targeting signals are the pieces of information that enable the cellular transport machinery to correctly position a protein inside or outside the cell.
This information is contained in the polypeptide chain or in the folded protein.
The continuous stretch of amino acid residues in the chain that enables targeting are called signal peptides or targeting peptides.
There are two types of targeting peptides, the pre-sequences and the internal targeting peptides.
The pre-sequences of the targeting peptide are often found at the N-terminal extension and is composed of between 6-136 basic and hydrophobic amino acids.
In case of peroxisomes the targeting sequence is on the C-terminal extension mostly.
Other signals are composed by parts which are separate in the primary sequence. To function these components have to come together on the protein surface by folding.
They are called signal patches.
In addition, protein modifications like glycosylation can induce targeting.

Protein translocation

In 1970, Günter Blobel conducted experiments on the translocation of proteins across membranes.
He was awarded the 1999 Nobel prize for his findings.
He discovered that many proteins have a signal sequence, that is, a short amino acid sequence at one end that functions like a postal code for the target organelle.
The translation of mRNA into protein by a ribosome takes place within the cytosol.
If the synthesized proteins “belong” in a different organelle, they can be transported there in either of two ways, depending on the protein.

Co-translational translocation

The N-terminal signal sequence of the protein is recognized by a signal recognition particle (SRP) while the protein is still being synthesized on the ribosome.
The synthesis pauses while the ribosome-protein complex is transferred to an SRP receptor on the endoplasmic reticulum (ER), a membrane-enclosed organelle.
There, the nascent protein is inserted into the translocation complex that passes through the ER membrane.
The signal sequence is immediately cleaved from the polypeptide once it has been translocated into the ER by signal peptidase in secretory proteins.
This signal sequence processing differs for some ER transmembrane proteins.
Within the ER, the protein is first covered by another protein to protect it from the high concentration of other proteins in the ER, giving it time to fold correctly.
Once folded, the protein is modified as needed (for example, by glycosylation), then transported to the Golgi apparatus for further processing and goes to its target organelles or is retained in the ER by various ER retention mechanisms.

Post translational translocation

Even though most proteins are co-translationally translocated, some are translated in the cytosol and later transported to their destination.
This occurs for proteins that go to a mitochondrion, a chloroplast, or a peroxisome (proteins that go to the latter have their signal sequence at the C terminus).
Also, proteins targeted for the nucleus are translocated post-translation.
They pass through the nuclear envelope via nuclear pores.

Sorting of proteins to mitochondria

Most mitochondrial proteins are synthesized as cytosolic precursors containing uptake peptide signals.
The pre-protein with pre-sequence targeted for the mitochondria is bound by receptors and the General Import Pore (GIP) (Receptors and GIP are collectively known as Translocase of Outer Membrane or TOM) at the outer membrane.
The pre-protein is translocated through TOM as hairpin loops.
The pre-protein is transported through the intermembrane space by small TIMs (which also acts as molecular chaperones) to the TIM23 or 22 (Translocase of Inner Membrane) at the inner membrane.
Within the matrix the targeting sequence is cleaved off by mtHsp70.
Three mitochondrial outer membrane receptors are known: TOM20, TOM22 and TOM70
The presequence translocase23 (TIM23) is localized to the mitochondrial inner membrane and acts a pore forming protein which binds precursor proteins with its N-terminal.
TIM23 acts a translocator for pre-proteins for the mitochondrial matrix, the inner mitochondrial membrane as well as for the intermembrane space.
TIM50 is bound to TIM23 at the inner mitochondrial site and found to bind pre-sequences.
Mitochondrial matrix targeting sequences are rich in positively charged amino acids and hydroxylated ones.
Proteins are targeted to sub-mitochondrial compartments by multiple signals and several pathways.
Targeting to the outer membrane, intermembrane space, and inner membrane often requires another signal sequence in addition to the matrix targeting sequence.

Sorting of proteins to chloroplasts

The pre-protein for chloroplasts contain a stromal import sequence or a stromal and thylakoid targeting sequence.
The majority of pre-proteins are translocated through the Toc and Tic complexes located within the chloroplast envelope.
In the stroma the stromal import sequence is cleaved off and intra-chloroplast sorting and folding continues.
Sorting of proteins to both chloroplasts and mitochondria
Many proteins are needed in both mitochondria and chloroplasts.
In general the targeting peptide is of intermediate character to the two specific ones.
The targeting peptides of these proteins have a high content of basic and hydrophobic amino acids, a low content of negatively charged amino acids.
They have a lower content of alanine and a higher content of leucine and phenylalanine.
The dual targeted proteins have a more hydrophobic targeting peptide than both mitochondrial and chloroplastic ones.

Protein destruction

Defective proteins are occasionally produced, or they may be damaged later, for example, by oxidative stress.
Damaged proteins can be recycled. Proteins can have very different half life, mainly depending on their N-terminal amino acid residue.
The recycling mechanism is mediated by ubiquitin.

MCQ’s

Q1. Which of the following organelles receives proteins via co-translational translocation?

[PYQ Type]
a) Nucleus
b) Mitochondria
c) Endoplasmic Reticulum ✅
d) Peroxisome
🧠 Explanation: Co-translational translocation means proteins are inserted into the ER during synthesis. This process is specific to the Endoplasmic Reticulum.

Q2. What is the role of the signal recognition particle (SRP) in protein targeting?

a) Degrades misfolded proteins
b) Assists in nuclear import
c) Binds the signal sequence and pauses translation ✅
d) Guides proteins to the lysosome
🧠 Explanation: SRP binds the N-terminal signal sequence and pauses translation, guiding the ribosome to the ER membrane.

Q3. Which signal is recognized by the M6P receptor during protein targeting?

a) Nuclear localization signal
b) Signal patch
c) Mannose-6-phosphate ✅
d) Stromal import sequence
🧠 Explanation: Lysosomal enzymes are tagged with Mannose-6-phosphate (M6P), which is recognized by M6P receptors for lysosomal targeting.

Q4. The TOM and TIM complexes are involved in the import of proteins into the:

a) Endoplasmic Reticulum
b) Chloroplast
c) Mitochondria ✅
d) Nucleus
🧠 Explanation: TOM (Translocase of Outer Membrane) and TIM (Translocase of Inner Membrane) help transport proteins into mitochondria.

Q5. Which organelle uses a C-terminal targeting sequence for protein import?

a) Nucleus
b) Peroxisome ✅
c) Chloroplast
d) Mitochondria
🧠 Explanation: Proteins destined for peroxisomes usually have a targeting sequence at the C-terminal end, often the SKL motif.

Q6. Which of the following organelles receives proteins via co-translational translocation?

a) Nucleus
b) Mitochondria
c) Endoplasmic Reticulum ✅
d) Peroxisome
🧠 Explanation: During co-translational translocation, proteins are synthesized directly into the ER.

Q7. Which organelle uses a C-terminal targeting sequence for protein import?

a) Nucleus
b) Peroxisome ✅
c) Chloroplast
d) Mitochondria
🧠 Explanation: Peroxisomes use a C-terminal SKL sequence for protein targeting.

Q8. The TOM and TIM complexes are involved in the import of proteins into the:

a) Endoplasmic Reticulum
b) Chloroplast
c) Mitochondria ✅
d) Nucleus
🧠 Explanation: TOM and TIM are translocase complexes that guide proteins into mitochondria.

Q9. What is the role of the signal recognition particle (SRP) in protein targeting?

a) Degrades misfolded proteins
b) Assists in nuclear import
c) Binds the signal sequence and pauses translation ✅
d) Guides proteins to the lysosome
🧠 Explanation: SRP recognizes the N-terminal signal, halts translation temporarily, and guides the complex to the ER.

Q10. Which signal is recognized by the M6P receptor during protein targeting?

a) Nuclear localization signal
b) Signal patch
c) Mannose-6-phosphate ✅
d) Stromal import sequence
🧠 Explanation: M6P tags lysosomal enzymes and is recognized by the M6P receptor for targeting.

Q11. Which of the following organelles receives proteins via co-translational translocation?

a) Nucleus
b) Mitochondria
c) Endoplasmic Reticulum ✅
d) Peroxisome
🧠 Explanation: Co-translational insertion occurs only at the ER, not in mitochondria or nucleus.

Q12. What is the role of the signal recognition particle (SRP) in protein targeting?

a) Degrades misfolded proteins
b) Assists in nuclear import
c) Binds the signal sequence and pauses translation ✅
d) Guides proteins to the lysosome
🧠 Explanation: SRP facilitates ER targeting by pausing translation and docking the ribosome.

Q13. The TOM and TIM complexes are involved in the import of proteins into the:

a) Endoplasmic Reticulum
b) Chloroplast
c) Mitochondria ✅
d) Nucleus
🧠 Explanation: These complexes transport proteins into and across mitochondrial membranes.

Q14. Which of the following organelles receives proteins via co-translational translocation?

a) Nucleus
b) Mitochondria
c) Endoplasmic Reticulum ✅
d) Peroxisome
🧠 Explanation: Only the ER allows insertion during protein synthesis.

Q15. What is the role of the signal recognition particle (SRP) in protein targeting?

a) Degrades misfolded proteins
b) Assists in nuclear import
c) Binds the signal sequence and pauses translation ✅
d) Guides proteins to the lysosome
🧠 Explanation: SRP binds the signal sequence and pauses translation to deliver to the ER membrane.

Q16. The TOM and TIM complexes are involved in the import of proteins into the:

a) Endoplasmic Reticulum
b) Chloroplast
c) Mitochondria ✅
d) Nucleus
🧠 Explanation: These complexes help proteins cross the outer and inner membranes of mitochondria.

Q17. What is the role of the signal recognition particle (SRP) in protein targeting?

a) Degrades misfolded proteins
b) Assists in nuclear import
c) Binds the signal sequence and pauses translation ✅
d) Guides proteins to the lysosome
🧠 Explanation: SRP identifies the N-terminal signal sequence and halts translation to guide the ribosome to the ER.

18. Which signal is recognized by the M6P receptor during protein targeting?

a) Nuclear localization signal
b) Signal patch
c) Mannose-6-phosphate ✅
d) Stromal import sequence
🧠 Explanation: M6P is a tag for lysosomal targeting, recognized by M6P receptors.

19. Which organelle uses a C-terminal targeting sequence for protein import?

a) Nucleus
b) Peroxisome ✅
c) Chloroplast
d) Mitochondria
🧠 Explanation: Peroxisomes typically use a C-terminal SKL motif (PTS1 signal) for targeting.

20.Which of the following is true about mitochondrial targeting signals?

a) They are rich in negatively charged amino acids
b) They are found in the C-terminal region
c) They are usually glycosylated
d) They are rich in positively charged and hydroxylated amino acids ✅
🧠 Explanation: Mitochondrial matrix-targeting sequences are positively charged and amphipathic, aiding recognition by TOM.

Q21. What is the function of the signal peptidase in protein sorting?

a) Degrades damaged proteins
b) Modifies glycoproteins
c) Removes signal peptides after translocation ✅
d) Adds M6P tags to lysosomal proteins
🧠 Explanation: Signal peptidase removes the signal peptide once the protein enters the ER lumen.

Q22. What characterizes a signal patch in a protein?

a) A continuous stretch of amino acids at the N-terminal
b) A glycosylation site
c) A folded structure bringing separate parts together ✅
d) An RNA sequence
🧠 Explanation: Signal patches form when separate parts of the protein come together after folding.

Q23. Which of the following protein import pathways is typically post-translational?

a) ER import
b) Mitochondrial import ✅
c) Golgi to lysosome
d) Co-translational targeting
🧠 Explanation: Proteins going to mitochondria, chloroplasts, and peroxisomes are generally imported post-translation.

Q24. The sequence ‘SKL’ at the C-terminal of a protein is a signal for targeting to:

a) Mitochondria
b) Nucleus
c) Peroxisome ✅
d) ER
🧠 Explanation: The Ser-Lys-Leu (SKL) motif is the PTS1 peroxisomal targeting signal.

Q25. Which receptor complex is involved in chloroplast protein import?

a) TIM
b) TOM
c) Toc and Tic ✅
d) SRP
🧠 Explanation: Toc (outer membrane) and Tic (inner membrane) complexes import proteins into chloroplasts.

Q26. Which proteins assist in the proper folding of proteins in the ER?

a) SRP
b) TIM
c) Chaperones ✅
d) TOM
🧠 Explanation: ER chaperones like BiP help in protein folding and prevent aggregation.

Q27. Which type of targeting signal must come together only after the protein has folded?

a) Pre-sequence
b) Signal patch ✅
c) Glycosylation site
d) Transit peptide
🧠 Explanation: Signal patches form after folding when distant residues spatially align to create a recognition site.

Q28. Which of the following is a transmembrane pore in the mitochondrial outer membrane?

a) TIM23
b) TOM40 ✅
c) SRP receptor
d) Tic110
🧠 Explanation: TOM40 forms the actual channel in the outer mitochondrial membrane for protein translocation.

Q29. What type of protein targeting occurs during protein synthesis?

a) Post-translational
b) Co-translational ✅
c) Nuclear localization
d) Glycosylation
🧠 Explanation: Co-translational targeting occurs as the protein is being synthesized and involves SRP and ER.