Endocytosis & Protein Degredation

Question 1

What are the 2 major routes of small volume endocytosis? How do they work?

Answer 1

Phagocytosis

• Multicellular organisms
• Usually by specialized cell (i.e. macrophage/ neutrophil)
• Cells recognize foreign organisms -> engulf-> deliver to lysosomes for degradation
• Recognize apoptotic cells

Pinocytosis
- Associated with vesicle uptake of (by) specific ligands and receptors

Question 2

What are the types of vesicles used in pinocytosis?

Answer 2

Clathrin coated &

Caveolae

Question 3

How are clathrin coated vesicles formed?

Answer 3

Cargo molecules (LDL) bind transmembrane receptor (LDLR) -> transmembrane receptor recognizes and binds adaptor protein (w/ motif in cytoplasm) -> Adaptor complex of proteins forms -> Enables clathrin coat to assemble on vesicle budding from plasma membrane (or golgi to secretory pathway)

Vesicle is pinched off from membrane by dynamin
- adaptor complex and clathrin rapidly dissociate

Question 4

What is the function of dynamin

Answer 4

Pinch clathrin coated vesicles off from golgi or plasma membrane

Question 5

What is the progression of phagocytosis of LDL?

Answer 5

LDL-receptor binding -> coated vesicle (clathrin and adaptor complex dissociation) -> early endosome -> late endosome (LDLR returns to membrane) -> lysosome

Question 6

What types of molecules enter through caveolae?

Answer 6

cholera, tetnus, folic acid,

Question 7

How many caveolins are in each vessicle?

Answer 7

144

Question 8

Where is caveolin 3 expressed? and what diseases can its mutation cause?

Answer 8

in skeletal and cardiac muscle

• limb girdle disese
• Rippling Muscle disease

Question 9

What can macrophages degrade?

Answer 9

EVERYTHING (except cholesterol)
mis-folded protiens
damaged organelles
good and bad endocytosed materials

Question 10

Why must improperly folded proteins be degraded?

Answer 10

They may aggregate and wreak HAVOC!

Question 11

What are the 3 major pathways of protein degradation?

Answer 11

Ubiquitin-proteasome system (UPS)
lysosome
autophagy

Question 12

What is autophagy used for?

Answer 12

• degradation of long lived proteins & entire organelles
• important in development
• Req for adaptation to environmental stresses (i.e. starvation)

Question 13

Describe the ubiquitin-proteasom system (UPS)

Answer 13

• Proteins deglycosylated after retrotranslocation out of ER
• Ligase enzymes E1, E2, and E3 bind & activate ubiquitin
• Polyubiquinated=> targeted to proteasome for degradation

Question 14

Describe the action of a proteasome

Answer 14

huge complex of proteis -> unwinds misfolded proteins, feed into compartment -> where cut into 7–9 amino acid lengths

Question 15

What % of cellular protein is proteasome?

Answer 15

~1%

Question 16

How many E1, E2, & E3 genes do humans have? What is the function of each?

Answer 16

E1: 1 - activate ubiquitin
E2: ~50 - Receive U from E1, transfer to E3
E3: ~500 - attaches U to lysine in protein - substrate specificity

Question 17

How many ubiquitins are required for degradation?

Answer 17

at least 4 in a chain (polyubiquitination)

Question 18

What are the other functions of ubiquitin?

Answer 18

multiubiquitination: mutiple single ubiquitins at several locations on a protein
aka: mono/multi ubiquitins- regulatory signals (u of histones, TFs to reg transcription

Question 19

What is the function of chaperone proteins?

Answer 19

Help with proper folding of newly synthesized proteins

Question 20

What are 2 examples of chaperone proteins?

Answer 20

hsp60
hsp70
(heat shock protein)

Question 21

How does hsp70 function?

Answer 21

prevents aggregation by binding to exposed hydrophobic areas in incompletely folded amino acids

Question 22

How does hsp60 function?

Answer 22

acts as an isolation chamber (prevent aggregation and refold) in barrel-shaped structure

Question 23

What is the generic name for proteins that bind sugas? Give 2 examples

Answer 23

lectins: calnexin & calreticulum

Question 24

What percentage of proteins are folded in the rER? Where are these proteins destined?

Answer 24

~30%

transmembrane proteins, vesicular transport and secretion

Question 25

Describe the protein folding pathway involving calnexin, calreticulum and a glucosyltransferase.

Answer 25

calnexin and calreticulum bind oligosaccharide chain if a glucose is present-> When glucose reomved by glucosidase (GT) C & C release protein -> if correctly folded allowed to exit ER

• > if incorrectly folded-> glucose added back on by glucosyltransferase and cycle repeats
• at a certian point (unclear when/why) protein is identified as unsaveable, “times out” and is translocated from ER, destined for degredation

Question 26

Describe a proteasome

Answer 26

~2million dalton protein
- central cylinder with cap a each end (recognize polyubiqutiinaiton) -> use ATP to unfold protein- feed into cylinder

Degrades only proteins!

Question 27

How many amino acids does ubiquitin have?

Answer 27

76 - HIGHLY CONSERVED!!

~1% of cellular protein

Question 28

Describe the function of a lysosome

Answer 28

• degrade extracellular materials& some intracellular components

Question 29

What ca lysosomes degrade

Answer 29

everything: proteins, lipids and sugars

Question 30

What is the path to a lysosome?

Answer 30

monoubiquinated plasma proteinstansferec by late endosome/ multivesicular body to lysosome
- multovesicular body (prelysosomal compartment with molecules slated for destruction in lumen)

lysosomal membrane proteins and transporters protected from degradation (error here = lysosomal storage disorder)

Question 31

What is autophagy?

Answer 31

when components of cell are broken down and reused

• normal turnover, damaged organelles, starvation
• uses double membrane around organelle- fuses with lysosome-> contents degraded

Question 32

What is ERp57

Answer 32

protein folding enzyme

thiol oxoreductase - allows formation of disulfide bonds

Question 33

What is UDP?

Answer 33

Folding sensor involved in quality control

glucose:glycoprotein glucosyltransferase