Name: sec62-sec63 pathway for er protein import
Uploaded: 2023-04-30T10:16:10.000Z
Duration: 1 min 27 s
Channel: JoVE

Some proteins are completely synthesized by free ribosome particles in the cytosol before entering the ER lumen. This mechanism is called post-translational protein translocation.
After translation, the cytosolic chaperones immediately bind to the polypeptide chain to prevent it from folding prematurely.
Once the chaperone-bound protein reaches the ER membrane, its signal sequence is recognized by a recognition site within the Sec61 channel.
Then, the protein slowly begins to glide down the channel and emerges at the luminal end, where it recruits ATP-bound BiP chaperone.
The heterodimeric Sec62/Sec63 complex hydrolyzes the ATP to induce a conformational change in the BiP molecule, allowing it to clasp the polypeptide.
New BiP molecules keep attaching to the incoming polypeptide, actively pulling it down into the ER lumen.
As the polypeptide enters the lumen, its signal sequence is cleaved off by the signal peptidase located in the ER membrane.
On completion of translocation, the nucleotide exchange factors replace all the ADPs bound to the BiP molecules with ATPs, preparing them for the next cycle of protein translocation.

post-translational translocation of proteins to the rer

Post-translational Translocation of Proteins to the RER

A sizable fraction of proteins destined for ER are first synthesized in the cell cytosol and then transported across the ER membrane&#8211;a process called post-translational translocation. Similar to cotranslationally translocated proteins, these proteins also use the Sec translocon complex to enter the ER lumen.
Targeting proteins to the ER
Hsp40 and Hsp70 chaperone molecules bind the translated proteins in the cytosol to prevent their folding. The chaperone binding helps to keep the signal sequence present on the target proteins available for interaction with the recognition site on the Sec61 channel.
The ER signal sequence of proteins translocating post-translation has characteristic features that distinguish them from cotranslationally translocated proteins. For example, in yeast, the signal sequence of post-translationally translocated proteins is relatively less hydrophobic compared to the signal sequence of cotranslationally translocating proteins. In contrast, in mammalian systems, the target proteins have a characteristic short positively charged N-terminal.
Translocon partners for post-translational translocation
All proteins enter the ER lumen using the Sec61 channel but the channel partners vary depending on whether it is a cotranslational or a post-translational translocation process. Sec62 and Sec63 play an important role during post-translational translocation. Sec62 has a strong affinity for the ER signal sequence present on the cytosol-translated proteins, while Sec63 has a specialized domain that hydrolyzes the ATP to allow the BiP chaperone to bind to the incoming protein. BiP chaperone is a lumenal chaperone that keeps the protein loosely folded or unfolded till it has attained the appropriate secondary structure. Together, the Sec62/63 complex and the BiP chaperone are involved in actively pulling the protein into the ER lumen by a Brownian ratcheting mechanism.
Other functions of the Sec proteins
Besides their central role in translocating proteins, Sec proteins are essential in other cell functions and diseases. For example,&#160; Sec62 is involved in the stress recovery of cells, while the Sec61 channel can double up as a passive ER calcium leak channel. Additionally, mutations and overexpression of the SEC genes are linked to diabetes, cancer, and various kidney and liver disorders in humans.

A sizable fraction of proteins destined for ER are first synthesized in the cell cytosol and then transported across the ER membrane&#8211;a process ...

post-translational-translocation-of-proteins-to-the-rer

Education

JoVE Core

Cell Biology

Unit 1

Transmembrane Transport in Endoplasmic Reticulum and Peroxisomes

transmembrane-transport-in-endoplasmic-reticulum-and-peroxisomes

KEY TERMS AND CONCEPTS

sec62-sec63 pathway for er protein import

Post-translational translocation moves some proteins from the cytosol into the endoplasmic reticulum (ER) after they are made. These proteins still use the Sec translocon complex to cross the ER membrane and enter the ER lumen. During this process, Hsp40 and Hsp70 chaperones bind the new protein in the cytosol and help stop it from folding too soon.

That chaperone binding also keeps the signal sequence exposed so it can contact the Sec61 channel. The ER signal sequence has features that help distinguish post-translationally translocated proteins from cotranslational ones. In yeast, the signal sequence is less hydrophobic than the one used by cotranslationally translocated proteins. In mammalian cells, the target proteins often have a short, positively charged N-terminal region.

Sec62 and Sec63 are important partners in this pathway. Sec62 binds strongly to the ER signal sequence on proteins made in the cytosol. Sec63 has a specialized domain that hydrolyzes ATP, which lets the BiP chaperone bind to the incoming protein.

BiP is a lumenal chaperone, meaning it works inside the ER. It keeps the protein loosely folded or unfolded until the right secondary structure forms. Together, the Sec62/63 complex and BiP help pull the protein into the ER lumen through a Brownian ratcheting mechanism.

Sec proteins also have other roles in cells. Sec62 helps cells recover from stress, and the Sec61 channel can act as a passive ER calcium leak channel. Changes in SEC genes, including mutations and overexpression, are linked to diabetes, cancer, and several kidney and liver disorders in humans.

Sec62-Sec63 Pathway for ER Protein Import

sec62-sec63-pathway-for-er-protein-import

the endoplasmic reticulum

ER Membranes, Microsomes, and Cell Support

The endoplasmic reticulum, or ER, is a major membrane network in eukaryotic cells. It makes up more than half of a cell’s membranes and about 10% of total cell volume. The ER also serves as a main site for protein and lipid synthesis for organelles such as the Golgi apparatus, lysosomes, secretory vesicles, and the plasma membrane. The ER was also the last major cell organelle to be discovered. In 1954, Keith Porter and George Palade produced the first high-resolution electron microscope...

smooth endoplasmic reticulum

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Smooth endoplasmic reticulum, or smooth ER, is a cell organelle with specialized jobs in both animal and plant cells. It is often linked to the tubular shape of the endoplasmic reticulum. Its main role is to support processes that need the right internal conditions inside the cell. Smooth ER is an important site for making steroid hormones and lipids. These lipids include phospholipids and triglycerides. It also makes major membrane lipids such as phosphatidylcholine, phosphatidylethanolamine,...

role of er in the secretory pathway

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The endoplasmic reticulum (ER) helps control protein secretion in eukaryotic cells. It acts as the first checkpoint in the secretory pathway, where many proteins are folded, modified, and tested for quality before they move on. The secretory pathway links membrane-bound compartments inside the cell and also connects cells to the outside environment. About one-third of the proteins made in a cell travel this route. These proteins move in membrane-bound vesicles between the ER, the ER-Golgi...

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directing-proteins-to-the-rough-endoplasmic-reticulum

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Proteins made in the cytosol often need a signal sequence to reach the rough endoplasmic reticulum (ER). These organelle-specific signals act like address tags and guide proteins to places such as the ER, mitochondria, and peroxisomes. Proteins sent to the ER may later move in vesicles through the Golgi complex to other organelles or leave the cell. The rough ER makes soluble proteins that can be sent to lysosomes or secreted outside the cell. It also makes transmembrane proteins, which become...

protein translocation machinery on the er membrane

protein-translocation-machinery-on-the-er-membrane

ER Membrane Translocon and Protein Entry

The translocon complex on the ER membrane acts as the main gateway for the protein secretory pathway. It moves new peptides into the ER lumen and also helps insert proteins into the ER membrane. In eukaryotes, the core of this complex is the Sec61 channel. Sec61 is a heterotrimer made of Sec61α, Sec61β, and Sec61γ, and it is the largest part of the translocon. In higher eukaryotes such as mammals, the channel is capped when it is resting. This helps prevent ions such as calcium from leaking...

cotranslational protein translocation

Sec61 and SRP in Protein Targeting

Sec61 and SRP help move newly made proteins across membranes during cotranslational translocation. This process is ancient and has been found in bacteria, archaebacteria, and eukaryotes. The machinery is conserved across these groups, which shows how important it is for cells. During cotranslational translocation, the Sec61 channel works with the signal recognition particle, or SRP, the SRP receptor, or SR, and ribosomes. SRP binds the ribosome through the polypeptide exit tunnel in the large...

Post-translational translocation moves some proteins from the cytosol into the endoplasmic reticulum (ER) after they are made. These proteins still use the Sec translocon complex to cross the ER membrane and enter the ER lumen. During this process, Hsp40 and Hsp70 chaperones bind the new protein in the cytosol and help stop it from folding too soon. That chaperone binding also keeps the signal sequence exposed so it can contact the Sec61 channel. The ER signal sequence has features that help...

insertion of single-pass transmembrane proteins in the rer

insertion-of-single-pass-transmembrane-proteins-in-the-rer

Single-pass Membrane Protein Topology in the ER

Single-pass membrane protein topology in the ER explains how certain proteins are built into a cell membrane. Integral membrane proteins are attached to the lipid bilayer of a cell organelle or membrane. They may span the membrane as transmembrane proteins, or they may sit on one side only as monotopic proteins. Transmembrane proteins have both membrane-spanning and extra-membrane domains. Their transmembrane regions usually contain 20-25 hydrophobic amino acids arranged in a helical shape.

insertion of multi-pass transmembrane proteins in the rer

insertion-of-multi-pass-transmembrane-proteins-in-the-rer

How Multipass Proteins Enter the ER Membrane

Multipass transmembrane proteins enter the rough ER membrane through a stepwise insertion process. These proteins are type IV integral membrane proteins, and they use multiple topogenic sequences to set their final position in the membrane. They can stay in the ER or later move to the Golgi complex, lysosome, or cell membrane. Most multipass proteins do not have a cleavable signal sequence. Instead, their first hydrophobic segment, also called a transmembrane domain, acts as the ER signal...

tail-anchoring of proteins in the er membrane

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ER Membrane Targeting of Tail-Anchored Proteins

Tail-anchored proteins are a group of membrane proteins that reach the endoplasmic reticulum, or ER, after they are made. They make up about 3% to 5% of membrane proteins in eukaryotic cells. These proteins have one transmembrane domain near the C-terminal end, which is the last part of the protein chain. Because the transmembrane segment is so close to the end, the signal recognition particle, or SRP, cannot bring these proteins to the ER for cotranslational insertion. Cotranslational...

gpi anchoring of proteins in the er membrane

gpi-anchoring-of-proteins-in-the-er-membrane

How GPI-Linked Proteins Reach the Membrane

GPI-linked proteins are made through a post-translational modification that helps place them on the outer side of the plasma membrane in eukaryotic cells. The GPI, or glycosylphosphatidylinositol, anchor is reversible and is built through 11 enzyme-driven steps. It includes a hydrophobic part and a hydrophilic part. The hydrophobic region contains phosphatidylinositol. The hydrophilic region includes phosphoethanolamine, glucosamine, mannose residues, and phosphate groups. Phosphoethanolamine...

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Rough ER protein folding changes help secretory and transmembrane proteins mature inside the ER lumen. These changes support folding and help stabilize the protein’s final three-dimensional shape. They happen at different stages as the protein folds. The main rough ER modifications fall into four groups: glycosylation, disulfide bond formation, assembly of protein subunits, and specific proteolytic cleavages, such as removal of signal sequences. N-linked glycosylation is the most common. In...

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export of misfolded proteins out of the er

export-of-misfolded-proteins-out-of-the-er

How the ER Clears Misfolded Proteins

The endoplasmic reticulum (ER) checks secretory and membrane proteins after they fold. Correctly folded proteins move through the calnexin cycle and continue to their final destination. Misfolded proteins are kept in the ER lumen, where ER chaperones try to unfold and refold them. When refolding fails, the protein becomes terminally misfolded. The ER then sends it to the cytosol through ER-associated degradation, or ERAD, for destruction. In this pathway, Hrd1 acts as both a ubiquitin ligase...

the unfolded protein response

ER Stress Control by the UPR

The endoplasmic reticulum, or ER, is a major site of protein synthesis in the cell. It also has strong quality control systems for protein folding and for breaking down misfolded proteins that cannot be fixed. Under normal conditions, some misfolded proteins are sent from the ER to the cytoplasm through ER-associated degradation, or ERAD. If ERAD cannot keep up, the cell turns on the unfolded protein response, or UPR. This response helps the cell adjust its protein folding pathways and reduce...

regulation of the unfolded protein response

regulation-of-the-unfolded-protein-response

IRE1 Signaling in ER Stress

IRE1 signaling helps cells respond to stress in the endoplasmic reticulum, or ER. IRE1, short for inositol-requiring kinase one, is the most conserved eukaryotic unfolded protein response receptor. It is a type I transmembrane protein kinase receptor with a site-specific RNase, or RNA-cutting, activity. The way misfolded proteins bind to the N-terminal domain of IRE1 is still not fully clear. Three activation models have been proposed: direct, indirect, and allosteric. Once a misfolded protein...

synthesis of phosphatidylcholine in the er membrane

synthesis-of-phosphatidylcholine-in-the-er-membrane

Phosphatidylcholine’s Roles in Cell Membranes

Phosphatidylcholine is a major glycerophospholipid in cell membranes. The endoplasmic reticulum, or ER, makes lipids that help build membranes and support cell functions such as energy storage and signaling. The lipid-making machinery sits in the ER membrane and gathers its reactants from the cytosol, the fluid inside the cell. After phosphatidylcholine and other lipids are made, the cell moves them to other organelles through the secretory pathway and through ER contact sites. Energy-rich...

assembly of the lipid bilayer in the er

How the ER Balances Membrane Lipids

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peroxisomes

Peroxisome Functions in Cells

Peroxisomes are specialized organelles found in fungi, plant, and animal cells. Their number, size, shape, and activity can change with the tissue type and the cell’s nutritional state. Cells with active lipid metabolism, such as adipocytes, neurons, and hepatocytes, often contain more peroxisomes than other cells. Peroxisomes help break down complex organic molecules. They also synthesize certain macromolecules and take part in redox signaling, which is a way cells use oxidation and reduction...

protein import into the peroxisomes

How Peroxisomes Receive Their Proteins

Peroxisomes receive many of their proteins from the cytoplasm and the endoplasmic reticulum. These membrane-bound organelles oxidize organic molecules by transferring hydrogen atoms to oxygen. This process produces hydrogen peroxide, which peroxisomes then break down into water and oxygen. Peroxisomes do not have the genetic machinery to make their own proteins. Most peroxisomal membrane, lumenal, and transmembrane proteins are made in the cytoplasm or the ER and then moved to the peroxisome.

key terms and concepts

See how Sec62, Sec63, and BiP move proteins into the ER after translation, and why ER signal sequences matter in high school cell biology.

Sec62-Sec63 Pathway for ER Protein Import | JoVE K12

cell biology

JoVE Core Cell Biology illustrates complex cellular processes in easy-to-understand yet comprehensive animated video lessons. This collection focuses on the essential components and functions of cells, including organelles and membranes, cellular transport, and cell energetics. It is complemented by scientist-in-action videos that demonstrate the application of cell biology in real-world research experiments.

Sec62-Sec63 Pathway for ER Protein Import

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