Protein Sorting II: Moving Proteins into Endomembrane System (L15)

Question 1

common pathway for protein sorting

Answer 1

1. protein synthesis and translocation across ER membrane
2. protein folding and modifications in the lumen of ER
3. protein transport to Golgi, lysosomes, or cell surface in vesicles

Question 2

how is trafficking done?

Answer 2

w/ vesicle packages - 50-100 nm in diameter that pinch off one compartment, then dock, fuse and drop off materials in next compartment

Question 3

what is the default pathway for packages?

Answer 3

secretion onto plasma membrane

Question 4

other functions of ER

Answer 4

• make steroids
• start point for cholesterol synthesis
• Ca sequestering
• cyt p450 detox
• lipid synthesis
• glycerophospholipid synthesis
• post translational modifications (glycosylation, disulfide formation, Pro or Lys hydroxylation)

Question 5

how are secreted proteins protected from degradation?

Answer 5

by being internalized in a membrane bound compartment

Question 6

composition of SRP

Answer 6

ribonucleoprotein complex of 6 proteins and a single 300-nt RNA molecule

Question 7

where does SRP bind?

Answer 7

• to SP on nascent polypeptide in cytoplasm and stops translation until docked on the ER
• also binds the large ribosomal subunit and the SRP-receptor on the ER

Question 8

SRP receptor

Answer 8

integral membrane protein, heterodimeric complex with a and B subunits

Question 9

function of SRP receptor

Answer 9

docks the SRP, ribosome, and nascent polypeptide onto the ER

Question 10

what is special about the SRP-p54 subunit and SRP receptor a subunit?

Answer 10

together, they can hydrolyze GTP (but cannot on own) to release SRP from ribosome and allow continued translation and translocation into ER

Question 11

what does BiP stand for? what is it?

Answer 11

binding protein - an Hsc 70 chaperone that uses ATP hydrolysis to draw in the protein

Question 12

what determines the topology of single-pass proteins?

Answer 12

internal stop-transfer-anchor (STA) and signal-anchor (SA) sequences

Question 13

when predicting which sides are cytosolic or luminal, where do the positively charged amino acids always stay?

Answer 13

positive residues always face cytosol

Question 14

what type of plot can help predict the type/class of integral membrane proteins?

Answer 14

hydropathy plots

Question 15

what does GPI stand for?

Answer 15

glycosylphosphatidylinositol

Question 16

what are GPI-anchored proteins?

Answer 16

a fifth type of membrane-anchored protein made from type I proteins by luminal transamidase

Question 17

function of GPI-anchored proteins

Answer 17

role in signaling, cell polarity, or interactions w/ ECM

Question 18

what post-translational protein modifications occur in the ER?

Answer 18

• proteolytic cleavages
• GPI-anchors
• N-linked glycosylation
• disulfide bond formation
• hydroxylation of Pro and Lys residues

Question 19

functions of ER

Answer 19

• post-translational protein modifications
• ensures proper protein folding
• assembly of multiprotein complexes
• targets misfolded proteins for degradation

Question 20

describe N-linked glycosylation of proteins in ER

Answer 20

• occurs on a subset of Asn residues
• adds on core of (GlcNAc)2(Man)3 always
• commonly adds on (Glc)3(Man)9(GlcNAc)2

Question 21

how is the mannose sugar tree for N-linked glycosylation

Answer 21

on a dolichol lipid anchor on the ER membrane

• add a phosphate +GlcNAc to dolichol phosphate
• add another GlcNac
• add 5 Man
• flip to ER lumen
• add 4 more Man
• add 3 glucose

Question 22

function of N-linked glycosylation

Answer 22

may promote folding and stability of glycoproteins

Question 23

where does disulfide bond formation occur? what protein does this?

Answer 23

formation and rearrangement occur in rER

catalyzed by protein disulfide isomerase (PDI)

Question 24

where does hemagglutinin formation occur? what proteins are responsible?

Answer 24

folding and assembly in ER

done by BiP, PDI, calnexin and calreticulin (lectins)

Question 25

describe unfolded protein response

Answer 25

1. increase in unfolded proteins causes BiP to dissociate to bind those proteins -> Ire1 makes dimer (endonuclease)
2. endonuclease dimer cleaves out unspliced intron of Hac1 mRNA to make a functional TF
3. spliced Hac1 mRNA translated
4. Hac1 TF taken into nucleus -> upregulates proteins necessary for proper protein folding

Question 26

what is the hereditary form of emphysema?

Answer 26

caused by point mutation in a1-antitrypsin -> misfolded protein -> elastase is not inhibited -> degrades fine tissue in lungs