Endoplasmic Reticulum and Protein Import - Vocabulary
Endoplasmic Reticulum (ER) – Comprehensive Study Notes
Context and onboarding
Exam coverage: early material included protein structure and function; new content will emphasize ER and secretory pathway.
Instructor emphasizes staying on top of ER material; prior exams included questions that were later dropped or revised after review.
Instructor offers to discuss exam questions and thought processes with students to reduce confusion.
ER basics and its place in the cell
ER stands for endoplasmic reticulum. It is the beginning of the secretory pathway and is intimately linked to the nucleus.
The outer membrane of the nucleus is continuous with the ER membranes (nuclear envelope).
Messenger RNA (mRNA) is transcribed in the nucleus, processed, and exits to the cytoplasm through nuclear pores to interact with ribosomes.
ER is the site where secreted and membrane proteins begin their journey through the secretory pathway.
ER is a major calcium storage site, important for signaling and binding site formation.
ER structure and topology
ER is highly folded; the membrane forms many cisternae that create a large surface area.
All the folds are connected as one continuous network (one big sack) rather than truly separate layers.
Terasaki ramps (parking garage analogy): interconnected levels/roofs of ER connected by ramps (Terasaki ramps) that level the movement of material between layers.
Terasaki ramps are named after the discoverer and are present at a precise angle (reported ~17°, not essential to memorize).
An interesting cross-disciplinary aside: a similar hexagonal/ladder-like structure observed in neutron star geometry; used to illustrate how similar structural motifs can appear in biology and physics, underscoring multiple realizations of efficient transport structures.
There are two ER types:
Rough ER (RER): studded with ribosomes on the cytosolic side, giving a rough appearance; site of importing proteins via the ribosome.
Smooth ER (SER): lacks ribosomes; major roles include lipid synthesis and calcium storage (calcium handling). In muscle tissue, a specialized SER is called the sarcoplasmic reticulum (SR).
The ER membrane is continuous with the outer nuclear membrane; ER membranes form the topologically distinct environment from the cytosol and lumen.
Key components of protein import into the ER
Targeting signals: proteins destined for the ER have an N-terminal signal peptide that is typically hydrophobic (about 15–30 amino acids long) with a hydrophobic core flanked by charged residues on one end.
Signal peptide cleavage: the signal peptide is usually cleaved after translocation by a signal peptidase, releasing the mature protein.
Nuclear-encoded proteins: all ER-destined proteins are translated by ribosomes, and the ribosome–polypeptide complex is targeted to the ER via the signal recognition particle (SRP).
Translation and ribosome types
Ribosome subunits: mammalian/eukaryotic ribosomes consist of a small 40S subunit and a large 60S subunit; the assembled ribosome is 80S during translation: 40S + 60S
ightarrow 80S.Ribosomes can be free in the cytosol or bound to the ER membrane (membrane-bound ribosomes on the rough ER).
Polyribosomes (polysomes): multiple ribosomes translate the same mRNA simultaneously to produce multiple copies of polypeptides; they can be attached to the ER membrane or free in the cytosol.
Polyribosome formation: typically forms around the nucleus where many ribosomes are present; can occur with both free ribosomes and those attached to the ER.
The signal recognition particle (SRP) and the SRP receptor (SR)
When the newly synthesized polypeptide contains an ER signal peptide, SRP binds the signal peptide as it emerges from the ribosome, halting translation.
SRP then recruits the ribosome–nascent chain complex to the ER membrane by binding to the SRP receptor (SR) on the ER membrane.
Both SRP and SR are GTPases; they bind GTP and hydrolyze it, providing energy to drive the targeting process and mounting the ribosome onto the translocon.
Upon GTP hydrolysis, SRP and SR dissociate, leaving the ribosome clamped to the translocon (Sec61) and ready for translocation.
Energetics recap:
SRP-SR interaction uses two GTP molecules; hydrolysis provides energy to clamp and align ribosome with the translocon.
Post-hydrolysis, SRP and SR recycle for another targeting cycle.
The Sec61 translocon and import mechanics
Sec61 is the protein-conducting channel (translocon) that forms a pore through the ER membrane.
The ribosome–nascent chain complex binds to Sec61; a signal peptide binds to a site inside the channel, and the rest of the polypeptide chain is threaded through Sec61 into the ER lumen or integrated into the ER membrane.
The translocon has a plug that blocks passage when not in use; when a ribosome binds and the nascent chain is translocating, the translocon opens to allow passage and then closes again when needed.
The signal peptide is cleaved by signal peptidase after translocation begins; once cleaved, the mature polypeptide begins to reside in the ER lumen or membrane.
The open state of Sec61 is triggered by ribosome binding; once a transmembrane domain is encountered, the channel can accommodate it and the domain slides into the membrane.
Hairpins and multiple transmembrane segments are possible; Sec61 can sequentially open and close to allow successive hydrophobic segments to exit laterally into the membrane.
Cotranslational vs post-translational import into the ER
Cotranslational import:
The polypeptide is threaded into the ER as it is being synthesized by the ribosome.
Energy source: GTP hydrolysis via SRP/SR and translation-associated processes; the ribosome translates with GTP-dependent steps, and Sec61 translocation is coordinated with translation.
Post-translational import:
The polypeptide is fully synthesized in the cytosol, often assisted by cytosolic chaperones (e.g., HSP70 family) to keep it unfolded and prevent premature folding.
The unfolded chain is then guided to the ER by chaperones (e.g., BiP in the ER lumen) and translocated through Sec61 using energy from ATP or associated ATPases in the lumen.
Energy sources across import modes:
Cotranslational: external GTP-driven energy (via SRP/SR) complements the translation process as the chain is fed into Sec61.
Post-translational: ATP-driven processes predominate (cytosolic HSP70s and BiP in the ER lumen; BiP uses ATP to pull the chain through Sec61).
Important caveat: Direct mitochondrial cotranslational import is not typical because mitochondria require crossing multiple membranes with different translocases; ER cotranslational import leverages Sec61 at a single membrane.
ER lumen chaperones and folding helpers
The ER lumen houses BiP (an HSP70 family chaperone) that helps pull the polypeptide into the lumen and prevents premature folding.
BiP provides an ATPase-driven mechanism to assist translocation and subsequent folding and quality control within the ER lumen.
The analogy to mitochondrial HSP70 (mtHSP70) and the related translocase complexes helps connect ER and mitochondrial import machinery (e.g., BiP vs mtHSP70, Sec61 vs analogous mitochondrial channel components).
Transmembrane protein insertion and topology rules
For proteins with transmembrane domains, the hydrophobic segments are integrated into the membrane as they are synthesized.
Orientation principles:
The plus end of a transmembrane domain tends to face the cytoplasm (positive-inside rule).
The minus end tends to face the lumen.
The terminal ends (N-terminus and C-terminus) relative to the membrane determine the final topology (which end is cytosolic vs luminal).
Single-pass transmembrane proteins:
A single hydrophobic segment traverses the membrane once and stops when the segment becomes embedded in the lipid bilayer.
Depending on where the charged residues sit, the remainder of the polypeptide will be synthesized on the cytosolic or luminal side.
Multi-pass (multipass) transmembrane proteins:
Contain multiple hydrophobic segments that form several transmembrane helices.
Each transmembrane segment may insert sequentially, forming “start” and “stop” signals that indicate where a segment begins and ends within Sec61.
The topological outcome is a polypeptide chain with alternating cytosolic and luminal loops, determined by the location of positive/negative charges around each hydrophobic segment.
Practical takeaway: When analyzing a multi-pass membrane protein, look for hydrophobic segments and the distribution of positively charged residues (plus/minus) to infer orientation (which side is cytosolic vs luminal).
Practical cellular context and implications
The ER is the entry point for proteins that will be secreted, integrated into membranes, or directed to the endomembrane system.
The calcium storage function of the ER is crucial for signaling and muscle contraction (and the sarcoplasmic reticulum in muscle tissue is a specialized form of SER involved in calcium handling).
The process of importing proteins into the ER is tightly coordinated with translation and relies on a defined set of players (signal peptide, SRP, SR, Sec61, BiP, etc.).
The secretory pathway includes subsequent trafficking steps from the ER to the Golgi and beyond, with proper folding, modification, and quality control.
Quick references to related topics and historical notes
Gunther Global proposed the signal hypothesis: the idea that a signal sequence directs ribosome-associated polypeptide chains to a translocation channel (Sec61) and that the signal peptide is cleaved during translocation, yielding a mature polypeptide in the ER lumen.
The classic diagram from Global’s work shows ribosomes on the ER surface, the translocon (Sec61), and the cleaving signal peptide, illustrating the cotranslational import process.
Ethical, philosophical, and real-world implications (as discussed in class)
An aside debated intelligent design: the observation that ER-like structures appear in biology and astrophysical contexts (e.g., Terasaki ramps vs neutron star structures) invites philosophical reflection about similarity, convergence, and interpretation of natural design.
Students are encouraged to draw their own conclusions and avoid over-generalizing from one structural analogy to universal claims about design.
Summary take-home points
The ER is the start of the secretory pathway, sitting at the interface between the cytosol and the ER lumen/membrane, and is continuous with the nuclear envelope.
There are two ER forms: rough (with ribosomes) and smooth (lipid synthesis, calcium storage; SR in muscle).
Protein import into the ER is initiated by an ER signal peptide, recognized by SRP, and delivered to the Sec61 translocon via the SRP receptor (SR).
Translation can be cotranslational (most common for ER-targeted proteins) or post-translational (in cases with strong hydrophobic segments and different chaperone requirements).
The process uses distinct energy currencies: GTP hydrolysis (SRP/SR and translation-linked steps) for cotranslational import and ATP (via BiP and HSP70 family) for post-translational import.
Transmembrane domain insertion follows a plus-inside rule; multiple transmembrane passes can yield complex topology, with hydrophobic segments forming helices that are integrated into the ER membrane.
The signal peptide is typically cleaved by signal peptidase; mature proteins are released into the ER lumen or embedded in the ER membrane as appropriate.
The ER’s architecture (with Terasaki ramps) supports efficient transport through a connected network, and ER calcium handling is central to many cellular processes.
Key terms to memorize
Endoplasmic reticulum (ER); rough ER (RER); smooth ER (SER); sarcoplasmic reticulum (SR); Terasaki ramps; translocon Sec61; signal peptide; SRP (signal recognition particle); SR (SRP receptor); BiP (ER HSP70); post-translational import; cotranslational import; polyribosome; hydrophobic signal/core; start/stop transmembrane domains; plus-inside rule; signal peptidase.
Practice cues for exams
Recognize the sequence of events in cotranslational import: ribosome synthesizes signal peptide → SRP binds → ribosome–SRP–mRNA complex binds SR on ER → GTP hydrolysis drives docking to Sec61 → signal peptide binds inside the channel → polypeptide translocates; signal peptide cleaved → mature protein in lumen or membrane.
Distinguish between RER and SER based on ribosome presence and functional roles.
Apply the plus-inside rule to predict orientation of multi-pass membrane proteins.
Note about visuals and terminology used in lectures
Electron micrographs of ER show stacked cisternae; approximated as “parking garage” with Terasaki ramps connecting layers.
Diagrams commonly depict Sec61 as a pore that opens upon ribosome docking; signal peptide moves into the channel and is cleaved.
The discussion includes a cross-disciplinary analogy to physics/astrophysics to illustrate universal transport principles and structural motifs across disciplines.
Quick glossary
SRP: Signal Recognition Particle that binds the ER signal peptide and halts translation to guide the ribosome to the ER.
SR: SRP Receptor on the ER membrane that interacts with SRP to dock the ribosome to Sec61.
Sec61: The translocon channel in the ER membrane through which polypeptides pass into the ER lumen or into the membrane.
BiP: ER-resident HSP70 chaperone that assists translocation and folding inside the ER lumen.
Pores and domains: Transmembrane segments are hydrophobic helices; their orientation is dictated by charged residues flanking the hydrophobic core.
References to details and numerical notes from lecture
ER signal peptide length: typically 15–30 amino acids; hydrophobic core central region.
The diameter/angle of Terasaki ramps in images is precise in some reconstructions (approx. 17°), but exact values are not essential for exam purposes.
The basic ribosome subunit sizes: 40S and 60S subunits join to form an 80S ribosome during translation.
Connections to foundational principles
Central dogma context: DNA → RNA → Protein; translation coupling with translocation into the ER shows how protein targeting is integrated with gene expression.
Membrane biology: topologically distinct environments (cytosol vs ER lumen) require specialized transport and folding mechanisms to maintain compartmental integrity.
Energy coupling: translation and translocation rely on nucleotide hydrolysis (GTP for SRP/SR and translation, ATP for BiP) to drive movement and folding.
Final takeaway
The ER orchestrates the beginning of the secretory pathway by recognizing ER-destined proteins, guiding them through Sec61, and directing them to the lumen or membrane with proper topology, while simultaneously supporting calcium storage and membrane biogenesis.