Biochem Block 4 - Week 2

Front 1

Which end of the peptide emerging from the ribosome contains the RER target sequence?

Back 1

N' end

Front 2

In the signal hypothesis, how does the peptide get into the RER? (6 steps)

Back 2

1. SRP: signal recognition particle binds to signal sequence of peptide and stalls translation

2. SRP binds SRP receptor in RER membrane, which is associated with a translocon

3. Binding of SRP to R mediates GTP binding

4. GTP hydrolysis opens translocon

5. SRP dissociates from ER R and ribosome

6. Peptide is translated through translocon

Front 3

What helps to push the growing peptide chain through the translocon pore from the cytosolic side?

Back 3

ATP hydrolysis during translational elongation

Front 4

What helps to push the growing peptide chain through the translocon pore from the ER lumen side?

Back 4

BiP

Sec63 complex

Front 5

What causes BiP to have a conformational change, binding peptide and racketing it through the translocon?

Back 5

Binding & hydrolysis of ATP

Front 6

In which type of membrane insertion is the signal cleaved?

Back 6

Type I

Front 7

In which types of membrane insertion is the C' end facing the cytosol?

Back 7

Types I and III

Front 8

How can proteins be anchored in the cytosolic side of the ER membrane?

Back 8

GPI linkage

GlycosylPhosphatidylInositol linkage

Front 9

On which side of the membrane are GIP-linked proteins linked?

Back 9

ER lumen, Extracellular leaflet

Front 10

What is the process of anchoring a single pass protein in the membrane?

Back 10

1. N' signal cleaved

2. Stop Transfer Anchor Sequence of hydrophobic AAs

3. Lateral peptide movement through translocon until STA is inside membrane

4. Remaining peptide is synthesized in cytosol

Front 11

The ++++ charged AAs next to the signal anchor sequence end up on which side of the ER membrane?

Back 11

Cytosolic side

Front 12

What is the anchor of a GPI-linked protein in the membrane? What is the end of the linker? Which end of the protein is linked to the GPI-anchor?

Back 12

Anchor: PI

End: phosphoethanolamine

C' end of protein is linked

Front 13

What enzyme is essential for creating GPI-anchors, cleaves proteins on the ER lumen side to attach the new C' to the anchor?

Back 13

GPI-transamidase

Front 14

What are 2 types of protein modifications that occur in the ER?

Back 14

1. Glycosylation

2. Disulfide bonds

Front 15

What are carbohydrate-binding proteins that can interact with other glycosylated proteins? Examples? (2)

Back 15

Lectins

ie Calnexin and Calreticulum

Front 16

What is the enzymatic addition of carbs to proteins?

The non-enzymatic addition?

Back 16

Glycosylation

Glycation

Front 17

In O-linked glycosylation: carbs are added to...

Back 17

-OH groups of serine and threonine

Front 18

In N-linked glycosylation: carbs are added to....

Back 18

asparagine

Front 19

What enzyme is involved in O-linked glycosylation?

In N-linked glycosylation?

Back 19

Glycosyl transferase

Oligosaccharyl transferase

Front 20

In N-linked glycosylation, glycans are added __________,

in O-linked, glycans are added __________

Back 20

N: as a preformed complex

O: sequentially (one at a time)

Front 21

What is an example of O-linked glycosylation?

Back 21

ABO blood groups

Front 22

What is the consensus sequence in N-linked glycosylation?

Back 22

Asn-X-Ser/Thr

Front 23

What is the backbone of the oligosaccharide complex that is used in N-glycosylation?

Back 23

Dolichol phosphate

Front 24

What is the first reaction in synthesis of the oligosaccharide complex that is used in N-glycosylation?

What inhibits this?

Back 24

UDP-GluNAc -> UMP

GluNAc phosphate is added to dolichol phosphate

Inhibits: tunicamycin

Front 25

In what form are sugar residues added to the oligosaccharide (for N-glycosylation)

Back 25

Activated NT-sugar donors (UDP-sugar or GDP-sugar)

Front 26

How is an N-glycosylation processed before the protein is ready for transfer to the Golgi?

Back 26

3 glucose and 1 mannose are removed

Front 27

Disulfide bonds are only found in which types of proteins?

Back 27

Secreted proteins or those in extracellular leaflet of plasma membrane

Front 28

What enzyme helps to produce disulfide bonds or rearrange them?

Back 28

PDI

(Protein Disulfide Isomerase)

Front 29

What enzyme helps restore PDI to its oxidized/disulfide bonded form?

Back 29

Ero1

Front 30

What protein deficiency causes liver disease in children and emphysema in adults?

What is the specific problem with that protein?

Back 30

alpha-1 antitrypsin

A point mutation prevents proper folding

Front 31

Anterograde proteins are which type of vesicle? What coat proteins?

What GTPase?

Back 31

COPII

Sec coat

Sar1 GTPase

Front 32

Retrograde vesicles are which type of vesicle?

What coat proteins?

What GTPase?

Back 32

COPI

Coatomer

ARF GTPase

Front 33

What proteins are associated with clathrin-coated vesicles? What GTPase?

Back 33

AP1, AP2, AP3, or GGA

ARF GTPase

Front 34

Where do clathrin-coated vesicles go?

Back 34

- trans-Golgi to endosomes or lysosomes

- PM to endosomes

Front 35

What is the general sequence of COPII vesicle formation? (4)

Back 35

1. COPII coat protein binds cytosolic domains of integral membrane cargo or R for soluble cargo

2. Curved COPII causes formation of vesicle, which pinches off ER

3. Vesicle de-coats, exposing v-SNAREs

4. v-SNAREs find t-SNAREs on cic-Golgi, vesicle fusion

Front 36

What ensures that ER resident proteins are retrograde transported back to the ER?

Back 36

KDEL sequence (Lys-Asp-Glu-Leu)

Front 37

What determines the loading and unloading of KDEL-proteins?

Back 37

pH!

pH in cis-Golgi is 6.6, but is 7.1 in ER

KDEL-proteins bind receptors at the lower pH, higher pH promotes release of KDEL proteins

Front 38

What is the purpose of the GTPase in vesicle formation?

Back 38

GTPase bound GTP: aids in binding of coat protein to target membrane

GTPase bound GDP: causes coat to disassemble to expose v-SNAREs

Front 39

What further processing of N-glycosylation occurs in the cis-Golgi?

Back 39

removal of three mannose sugars i.e. High Mannose Glycosylation

Front 40

At what place in the Golgi does High Mannose glycosylation occur? How about Complex Glycosylation?

Back 40

Mannose: after cis

Complex: after trans

Front 41

How does the AP assist in getting the proper cargo to the appropriate type of transport?

Back 41

It binds the sorting signal on cargo and interacts with the coat protein (usually clathrin)

Front 42

Transport to lysosome from TGN involves:

- which AP?

- which GTPase?

- which cargo signal?

Back 42

AP3

ARF
Mannose 6-phosphate

Front 43

Transport to endosome from TGN involves:

- which AP?

- which GTPase?

- which cargo signal?

Back 43

AP1

ARF

Tyr-X-X-bulky hydrophobic

Front 44

Transport from plasma membrane to endosome involves:

- which AP?

- which GTPase?

Back 44

AP2

ARF

Front 45

In regulated secretion of vesicles, cargo forms internal aggregates involving.... (3)

Back 45

Chromogranin A, Chromogranin B, Secretogranin II

Front 46

Clathrin protein is composed of?

Back 46

3 heavy & 3 light chains

Front 47

What is the 2-step process that occurs within the cis-golgi to create the mannose 6-phosphate signal?

Back 47

1. UDP-GlcNAc is used to add GluNAc-phosphate to one of the mannose sugars (enzyme: GlcNAc-1-phosphotransferase)

2. GluNAc is then removed (enzyme: phosphodiesterase)

Front 48

What enzyme is defective in Inclusion Cell (I-Cell) disease? What is the result of this?

Back 48

GlcNAc phosphotransferase is defective

No M6P labeling of lysosomal enzymes

Front 49

What is the less-severe form of I-Cell disease?

Back 49

pseudo-Hurler polydystrophy

Front 50

What is the classification of I-Cell Disease?

pseudo-Hurler?

Back 50

I-Cell: Mucolipidosis II

pseudo-Hurler: ML III

Front 51

What are the symptoms of I-Cell disease?

Back 51

Coarse features

Child reaches milestones, but then regresses

Psychomotor retardation, below average growth rate

Front 52

What is the pH of a lysosome?

Back 52

5-6

Front 53

Cytosolic proteins containing what sequence can be selectively taken up by lysosomes?

Back 53

KFERQ

Front 54

What GTPase is involved in the budding of a clathrin coated endocytotic vesicle?

Back 54

Dynamin

Front 55

What protein promotes de-coating of the clathrin coat on an endocytotic vesicle?

Back 55

ARF

Front 56

What are the different domains of the LDL receptor?

Back 56

1. Extracellular, ligand-binding domain

2. Extracellular, beta-propeller domain

3. Cytosolic surface, NPYX sorting seq

Front 57

What causes release of the LDL particle within the lysosome?

Back 57

- Beta-propeller domain becomes positively charged in the acidic environment, and binds tightly to the ligand-binding domain

- this interferes with interactions with ApoB, so the LDL is released

Front 58

What is the molecular basis for familial hypercholesterolemia?

Back 58

Mutation in LDL receptor

Front 59

Ferrotransferrin has high affinity for _______ at ______ pH

Back 59

High affinity for transferrin R at neutral pH (extracellular matrix)

Front 60

What promotes release of iron from ferrotransferrin?

Back 60

High acidity

Front 61

Apotransferrin has high affinity for the transferrin receptor at ______ pH and low affinity for the receptor at ______ pH

Back 61

high affinity at low pH

low affinity at neutral pH

Front 62

What proteins and protein modifications are critical to invagination of membrane for degradation in the lysosome?

Back 62

1. Hrs proteins (monoubiquitinated)

2. ESCRT complex

Front 63

Which HIV protein is analogous to Hrs and allows the HIV to assemble ESCRT and bud?

Back 63

HIV Gag protein