Membranes L2

Question 1

How can you investigate cell fusion?

Answer 1

Make antibodies human and mouse proteins
Add dye that fluoresces green under UV light 4 human
Red for mouse
Treat cells w antibodies and fuse cells
After 40 mins fluorescence intermixed ie. proteins move in membrane

Question 2

what is FRAP and what do results of this experiment show?

Answer 2

Fluorescence Recovery After Photobleaching (see diagram in notes)

Question 3

what does FRAP experiment reveal?

Answer 3

45% glycoproteins immobile, 55% mobile

Question 4

Describe erythrocycte cytoskeleton

Answer 4

see diagram in notes

Peripheral proteins lie beneath plasma membrane in cytosolic component and form cytoskeleton

Question 5

What happens when erythrocytes are treated w low ionic strength buffers?

Answer 5

spectrin and actin dissociate from membrane

shape lost, lateral mobility of membrane proteins observed

Question 6

What determines membrane protein fate when its being translated?

Answer 6

signal sequence at N terminus (At 5’ end, first bit to be translated)

Question 7

Describe process of protein being put in ER

Answer 7

See notes

Question 8

Function of foldases?

Answer 8

bind exposed hydrophobic surfaces in ER that should be buried in fully folded protein
Their reactions are ATP-dependent

Question 9

How does protein get fixed in bilayer?

Answer 9

‘Stop-transfer’ sequences stops protein moving through pore

Question 10

signal peptidases…

Answer 10

cleave N-terminus signal sequences

Question 11

What are retention sequences

Answer 11

Sequences that show protein destined to stay in ER, not exported

Question 12

Sorting sequences…

Answer 12

Target proteins to cellular compartments

Question 13

Describe glycosylation

Answer 13

Sugars making up main structure added in reaction catalysed by glycosyl transferases during passage of protein into ER
Side chains added during passage through Golgi Apparatus

Question 14

Describe process of endocytosis (up to vesicle uncoating)

Answer 14

1) solute to be transported binds extracellular domain of receptor
2) In C terminus of receptor is specific tetrapeptide sequence allowing receptor to bind protein adaptor
3) Protein adaptor interacts with cytoplasmic protein clathrin
4) Clathrin assembles beneath PM and polymerises to form coated pits of invaginated membrane
5) Invagination continues to give clathrin-coated vesicle
NB: if essay asks about transport ACROSS membrane don’t talk about exo/endocytosis

Question 15

How is extracellular cholesterol transported cell-to-cell

Answer 15

Its insoluble and transported as low-density lipoprotein (LDL) , a core of cholesterol ester-linked to fatty acids surrounded by monolayer of phospholipids and unesterified cholesterol

Question 16

LDL receptor…

Answer 16

Is a single span glycoprotein in the PM, recognises specific protein in monolayer

Question 17

What is the CURL

Answer 17

The Compartment of Uncoupling of Receptor and Ligand, fuses with cholesterol vesicle once uncoated, has low pH of 5.5 which causes dissociation of LDL from receptor

Question 18

What happens to LDL after CURL?

Answer 18

Gets transported by vesicles to lysosome containing hydrolytic enzymes to cleave fatty acids from cholesterol

Question 19

Draw clathrin

Answer 19

(see notes for diagram)