Liu

Question 0

Where are Ran-GAP and Ran-GEF located and what do they do?

Answer 0

Ran-GAP: cytosol, hydrolysis of Ran-GTP to Ran-GDP

Ran-GEF: nucleus, dissociation of GDP from Ran-GDP, which is quickly exchanged for GTP

Question 1

A signal sequence is an amino acid chain that directs a protein to a certain organelle. What is the 3-D structure made up of multiple internal amino acid sequences known as?

Answer 1

signal patch

Question 2

Proteins destined for the mitochondria have an N-terminal signal/leader sequence with many _____ amino acids. They are transported across the outer membrane by ____ proteins and across the inner membrane by ____ proteins.

Answer 2

positively charged (K, R); TOM complex, TIM complex

Question 3

Transport into ____ occurs co-translationally, whereas transport into ____, ____, and ____ occurs post-translationally.

Answer 3

ER; mitochondria, nuclei, peroxisomes

Question 4

Proteins destined for ____ have a specific 3-amino acid import sequence on their ___ terminus.

Answer 4

peroxisomes, C-terminus

*Even large oligomeric proteins do NOT have to be unfolded for transport to peroxisomes.

Question 5

Transport into ____ requires fully synthesized but unfolded peptides and the use of chaperones and ATP.

Answer 5

mitochondria

Question 6

What is a microsome?

Answer 6

small ER particle resulting from homogenization of cells, either a rough microsome (RER) or a smooth microsome (SER). Rough microsomes are denser due to their ribosome content.

Question 7

How do SRPs help translocate proteins to the ER?

Answer 7

Signal recognition particles (SRPs) are proteins that bind the N-terminal signal sequence of the protein being imported and the ribosome itself, causing a pause in translation. The ribosome-SRP complex binds the SRP receptor on the ER membrane, and the nascent peptide is fed through a nearby Sec-61 translocator complex. The SRP dissociates and translation continues concurrent with translocation.

Question 8

For soluble proteins, ER signal sequences are recognized twice. Why? Comment on their hydrophobicity.

Answer 8

Signal sequences are hydrophobic and are bound by hydrophobic signal-binding domains of the SRP, which brings the ribosome/nascent peptide to the SRP receptor on the ER membrane. Then they bind the Sec61 translocator complex at the hydrophobic start-transfer-peptide-binding site, which opens the pore and allows translocation of the soluble peptide.

Question 9

How are transmembrane proteins embedded in the ER membrane?

Answer 9

Start-signal binding at start-signal-peptide-binding site, translation and translocation, then stop-signal binding at stop-signal-peptide-binding site, and termination of translocation fixing the peptide to the membrane and leaving a cytoplasmic domain in the cytosol.

Question 10

A mutation in which protein of the ER-associated degradation pathway causes autosomal recessive juvenile Parkinson’s (AR-JP)? What is its substrate?

Answer 10

Parkin (an E3 ubiquitin ligase); Pael receptor (a transmembrane protein normally degraded by Parkin that accumulates in AR-JP and damages the ER)

Question 11

What are the 3 major classes of glycoproteins (based on the nature of the linkage between their polypeptide chains and their oligosaccharide chains)?

Answer 11

1. O-linked: Glycosidic bond between Ser/Thr OH and a sugar such as N-acetylgalactosamine (GalNAc-Ser[Thr])
2. N-linked: Glycosidic bond between amide nitrogen of Asn and N-acetylglucosamine (GlcNAc-Asn)
3. GPI-anchored: C-terminal amino acid joined to an oligosaccharide (via a phosphorylethanolamine moiety), which in turn is linked to phosphatidylinositol (via glucosamine).

Question 12

What are the three subtypes of N-linked glycoproteins? What do they all have in common? How is the oligosaccharide added to the polypeptide?

Answer 12

complex, hybrid, high mannose; pentasaccharide core Man3(GlcNAc)2; the oligosacch. group is transferred from a Dolichol-P-P-oligosaccharide complex to the Asn of the peptide chain using oligosaccharide transferase whenever the sequence Asn-X-Thr/Ser occurs

Question 13

Which drug inhibits the formation of the dolichol-P-P-oligosaccharide complex necessary for N-linked glycosylation?

Answer 13

tunicamycin

Question 14

The GPI anchor is assembled independently and then transferred to the ______ end of its acceptor protein, thereby cleaving the transmembrane acceptor protein on the ____ side of the membrane, leaving a small hydrophobic peptide within the membrane.

Answer 14

C-terminal, ER lumen

Question 15

What are the 3 major types of vesicle coats and what route do their vesicles primarily take?

Answer 15

COP I: Golgi –> ER (mostly)
COP II: ER –> Golgi
Clathrin: Membrane Golgi

Question 16

Describe the structural framework of a clathrin coat.

Answer 16

Clathrin is the major coat protein that provides the mechanic force needed for budding and is the framework of the coat.

3 light chains and 3 heavy chains -> 3-legged triskelion -> clathrin cage

Adaptins - coat proteins that link clathrin cage with transmembrane receptor

Question 17

What are adaptins?

Answer 17

Coat proteins that link clathrin with various transmembrane receptors. There are at least 4 types of adaptins, each specific for a different set of cargo receptors.

clathrin–adaptin–cytoplasmic domain of transmembrane receptor–membrane–extracellular domain–cargo

Question 19

Which protein is used in endocytotic vesicle budding?

Answer 19

Dynamin creates a collar around the vesicle and pinches it off, clathrin coat quickly dissociates

Question 20

Small GTPases are involved in the recruitment of coat proteins from the cytosol to the different membranes (Golgi, ER, plasma). ____ is responsible for ____ and ____ coat assembly at Golgi membranes. _____ is responsible for _____ coat assembly at the ER membrane.

Answer 20

ARF, COP-I, clathrin

Sar1, COP-II

Question 22

When vesicle encounters the correct target membrane, the binding of ____ to _____ causes the vesicle to remain bound for long enough to allow ___ to hydrolyze its GTP, which locks the vesicle onto the target membrane (“docking”). These, together with ___ induce the two membranes to fuse. A similar fusion process is employed in _____ receptors.

Answer 22

t-SNARE, v-SNARE, Rab-GTP; NSF; chemokine

Question 23

Resident ER proteins are retrieved by ____ receptors. They are packaged in ____-coated vesicles for retrograde transport back to the ER.

Answer 23

KDEL (recognize K-D-E-L sequence on proteins), COP-I

Question 24

What happens to proteins secreted from the ER that don’t have a KDEL sequence? KDEL sequence at C terminus?

Answer 24

not retrieved (cytosol?); stay in ER

Question 25

What is the”default” secretory pathway, constitutive or regulated?

Answer 25

constitutive

Question 26

What are the 3 best-understood pathways of protein sorting in the Golgi?

Answer 26

constitutive secretion, regulated secretion, diversion to lysosomes

Question 27

Docking of neurotransmitter vesicles to the presynaptic membrane is similar to vesicle transport between ER and the Golgi, using ___ and ____ (v-SNARE and t-SNARE analogs), and the synaptic vesicle specific G-protein _____.

Answer 27

synaptogamin, syntaxin, Rab3

Question 28

What is the difference between nonselective and selective pinocytosis?

Answer 28

nonselective: cell ingests whatever solutes are in the extracellular space
selective: receptor-mediated, selectively ingests certain solutes at a higher concentration because they bind to receptors

both use clathrin-coated pits

Question 29

Although clathrin-coated pits make up only __% of the plasma membrane surface, ___% of LDL receptors are localized in them. What does it mean that ‘LDL receptor uptake is a continuous process’?

Answer 29

2%, 50-80%, endocytosis of LDLR occurs with or without LDL bound to it.

Question 30

In familial hypercholesterolemia, there are at least 5 different LDLR mutations that can lead to the disease. Which class of LDLR mutations prevents aggregation of LDLR in the coated pits? What fails?

Answer 30

C-terminal cytoplasmic domain mutations; LDLR still binds LDL but doesn’t aggregate in coated pits because of a mutation in the adaptin-binding signal sequence FRXY. This cytoplasmic tail normally guides the receptor to the coated pit by recognizing and binding adaptin (Remember, adaptin is the link between the clathrin cage and other proteins). FRXY uses a tyrosine in a tight turn (seems to be important)

Question 31

What are the 3 outcomes of receptor-mediated endocytosis?

Answer 31

Recycling: receptor is recycled back to the membrane
Degradation: receptor and ligand are degraded
Transcytosis: receptor and ligand are transferred from extracellular space on one side of a polarized cell (e.g. simple epithelium) to extracellular space on opposite side (i.e. apical to basal)

Question 32

Describe how newborn mammals get IgG into their bodily tissues.

Answer 32

IgG-receptor-binding is pH-sensitive. The baby gets IgG from the mother’s milk. Milk -> intestinal lumen, where IgG binds IgG-R at pH 6, then transcytoses across intestinal epithelium, enters extracellular fluid in circulation with pH 7 which causes IgG to dissociate from its receptor.

Question 33

What is different between EGFR and LDLR accumulation in coated pits?

Answer 33

LDLRs accumulate with or without LDL. EGFRs accumulate in coated pits only after binding EGF.

*Also, after they unload their contents in the cell, LDLRs are recycled and EGFRs are degraded.

Question 34

How are receptors degraded?

Answer 34

Tagged with ubiquitin, transferred to endosome so ubiquitin tag is on the cytoplasmic side, endosome creates pseudopodia-like extensions and envelops the ubiquitin-receptor complex bound to it, forming a multivesicular body. It then fuses with lysosomal vesicles that contain degradative enzymes.

Question 35

When a lysosome uses all of its enzymes it is called a _________.

Answer 35

residual body

Question 36

Lysosomal proteins are synthesized in the ___ and transported through the Golgi. In the lumen of the cis-Golgi, _______ is added exclusively to _______ on the proteins. They are then transported from the TGN to a _______, eventually forming a _____.

Answer 36

RER, mannose-6-phosphate, N-linked oligosaccharides, late endosome, lysosome

Question 37

What is Inclusion-cell disease?

Answer 37

Lack of the first enzyme in lysosomal protein sorting, GlcNAc-phosphotransferase (no P in M6P). Failure to properly signal lysosomal enzymes results in secretion of enzymes to the blood. This doesn’t cause much of a problem because most hydrolases are neutralized at blood pH, but now lysosomes have no degradative enzymes and things start accumulating in the cell as inclusions.