Endomembrane System Part 3 Flashcards
How do the membranes form?
Membranes do not form de novo - all membranes arise from pre-existing membranes
Where are membrane proteins and lipids synthesized?
Most are synthesized at the ER
Glycolipids synthesized in the Golgi and unique chloroplast and mitochondrial proteins & lipids
Where do ER membrane proteins and lipids traffic?
Nascent ER membrane proteins & lipids can traffic to other membranes in the cell
For example, move to other ER subdomains (via lateral diffusion through the bilayer) OR to other ‘downstream’ organelles of the endomembrane system (via transport vesicles)
Results in each organelle possessing a unique complement of membrane proteins & lipids
Where are ER membrane proteins and lipids distributed?
Nascent ER membrane proteins and lipids are distributed and/or oriented in the lipid bilayer in an asymmetric manner
What are the three types of membrane proteins and what they do?
Integral Membrane Proteins: Different regions of protein located on either the cytoplasmic or exoplasmic (i.e., ER lumen) face of the ER membrane
Peripheral Membrane Proteins: Located on either the cytoplasmic or luminal side of the ER membrane
Membrane phospholipids: Distributed unequally between the cytoplasmic and exoplasmic leaflets of the ER membrane bilayer
Why does membrane topology need to be the right way?
Needs to be set up the right way to ensure the correct protein function
How is protein and lipid asymmetry established?
It is established at the ER and maintained throughout the rest of the endomembrane system, the orientation remains the same
Cytoplasmic and endoplasmic faces of cellular membranes are conserved in the endomembrane system
ER luminal protein or region(s) of ER membrane-spanning protein facing the lumen are located in the lumens of all other endomembrane compartments in which it resides or at the plasma membrane, on the exterior of the cell
How are newly synthesized proteins in the ER processed?
The final steps in the co-translation translocation pathway involve ‘processing’ of the newly synthesized protein in the ER lumen
1) Signal sequence cleavage: removal of the N-terminal signal sequence by signal peptidase
2) Initial stages of glycosylation: covalent addition of unique carbohydrate side chains to specific amino acids of the nascent protein (required for proper folding, protein-protein binding, etc) –> Add sugar groups to help determine if folded directly
3) Protein folding and assembly: nascent protein is folded into the proper 3D conformation and oligomeric assembly by molecular chaperones (reticuloplasmins)
4) Quality Control: Misfolded and/or improperly assembled proteins are recognized and degraded
ER serves as ideal processing and quality control site for nascent proteins since it represents the first compartment in the endomembrane system (biosynthetic & secretory pathway)
Explain glycosylation
Most proteins (soluble and membrane) synthesized in the ER are glycoproteins (have sugar groups added) Glycoproteins are proteins linked to one or more sugar chain (oligosaccharides) attached to specific amino acids within the nascent polypeptide Sugar groups aid in the protein's proper folding AND serve as binding sites for other macromolecules that interact with the protein. Glycosylation ensures proper folding of a protein
What is N-linked glycosylation?
It is the most common type of glycosylation
The addition of specific short chains of sugar monomers (linked together in a specific order to form an oligosaccharide) to the terminal amino group of asparagine (N)
What does N-linked glycosylation consist of?
Two stages:
i) Core glycosylation
ii) Core modification
For some glycoproteins that are transported to other post ER compartments, the core modification stage continues in the Golgi
What is Core glycosylation?
The first stage of N-linked glycosylation
Various ER membrane-bound glycosyltransferases synthesize the core oligosaccharide –> highly branched oligosaccharide chain consisting of 14 sugar residues, including a 3-glucose long terminal branch (important for quality control)
The process begins with the addition of the first sugar to dolichol phosphate - membrane lipid serving as an ‘anchor’ and ‘carrier; for the new, growing core oligosaccharide
What does Tunicamycin do?
Blocks the first step of N-linked glycosylation (inhibits glycosyl-transferase action), preventing the subsequent proper folding of nascent ER proteins
What are the steps of core glycosylation?
Glycosyltransferases continue to add (one at a time) sugar units at specific positions on the growing core oligosaccharide Synthesis of core oligosaccharide begins on the cytoplasmic face of the ER membrane and ends on the ER luminal (exoplasmic) face --> core oligosaccharide precursor is 'flipped' across the ER membrane during its synthesis Final sugars (mannose & glucose units) added via 'flipping' carrier dolichol phosphates The final step involves the transfer of core oligosaccharide from dolichol lipid carrier to nascent soluble or integral membrane protein while still being synthesized via the Sec61 co-translocational translocation pathway --> empty dolichol carrier recycled (& flipped) for another round of core oligosaccharide synthesis --> Core only transferred to the luminal-facing portions of nascent ER proteins with a specific amino acid sequence motif: N-x-S/T (core attached to asparagine residue)
What is core modification?
It is the second stage of N-linked glycosylation
What are the core modification steps?
After transfer to the nascent protein, the 14 sugar oligosaccharide(s) is gradually trimmed and modified
Two (of the three) terminal glucose units are removed (‘trimmed’) by ER lumen glucosidases
Subsequent removal (and re-addition) of the last glucose unit is important for the process that ensures proper protein folding/assembly (i.e, quality control)
During N linked glycosylation and modification, the nascent protein is rapidly folded into the proper 3D conformation which is mediated by several ER lumen and membrane proteins
Core oligosaccharide(s) added to nascent protein during N-linked glycosylation also contribute to proper protein folding/assembly and stability and participate in protein quality control
The addition of sugar and all listed above ensure proper folding
What is reticuloplasmins?
ER molecular chaperones, including BiP, calreticulin, and calnexin
Bind transiently (reversibly) to nascent ER proteins to prevent misfolding or aggregration
Involved with core-modification
BiP = Binding Immunoglobin Protein
What is protein disulfide isomerase (PDI)?
The enzyme catalyzes the formation of intra/intermolecular disulfide bonds
The disulfide bonds between cysteine residues on the same or different (nascent) polypeptides promote proper folding and assembly by stabilizing their proper 3D conformation
Involved with core-modification
