Phosphatidylserine and Blood Coagulation

Question 1

how is the loss of blood stopped after a cut

Answer 1

Forming a blood clot at the damaged area

Question 2

What is a blood clot? When do they form?

Answer 2

Plug of platelets, red blood cells, and fibrin molecules. Blood clot forms from the action of a cascade of extracellular reactions (clotting factors which convert fibrinogen to fibrin)

Question 3

What are platelets?

Answer 3

Key role in clot formation
Disk shaped cells with no nucleus
smaller than red blood cells
Gather and aggregate at sites of blood vessel damage
Go from resting to activated which activates the blood coagulation cascade

Question 4

Where does the blood coagulation cascade occur? When are the proteases activated?

Answer 4

Occurs on the exterior surface of platelets (extracellular)
requires the activation of coagulation factors present in the blood
Proteases are activated in response to blood vessel damage
Activation of one protease is achieved by its proteolysis by another

Question 5

T/F most coagulation factors are active proteases

Answer 5

False, most are inactive proteases

Question 6

Intrinsic vs extrinsic pathway for blood coagulation cascade

Answer 6

Intrinsic: internal damage to blood vessel wall (high blood pressure)

Extrinsic: External trauma (cut finger with a knife)

Question 7

What is the synonym for prothrombrin and fibrinogen

Answer 7

Factor II - prothrombrin
Fibrinogen- Factor I

Question 8

What is the blood coagulation cascade

Answer 8

Coagulation cascade activates factor X to factor Xa
Factor Xa combines with factor V and PS to cleave prothrombin and activates it to thrombin
Thrombin activates fibrinogen to fibrin
Fibrin forms clots

Question 9

What is factor X

Answer 9

Part of the prothrombinase complex

Question 10

What is the prothrombinase complex made of? What does the prothrombinase complex do?

Answer 10

Made of Factor X, Factor V, and phosphatidylserine
Converts prothrombin to thrombin
Xa and V interact and bind to PS on the surface of a platelet

Question 11

How does binding to PS and Factor V affect the Km and Vmax of Xa for prothrombin

Answer 11

Decreased Km and increased Vmax

increased rate of reaction

Question 12

What is the activity of factor Xa before forming the prothrombinase complex

Answer 12

It is active but activity is low unless it interacts with Factor V/PS

Question 13

What molecule is coagulation dependent on? Is there a problem with that

Answer 13

Coagulation depends on PS present on the platelet
surface, but most of it is in the inner leaflet of the
plasma membrane

There is not enough PS in the outer leaflet of the platelet membrane for prothrombinase complex to function

Question 14

How is phospholipid asymmetry of the plasma membrane of platelets achieved? Does this require energy?

Answer 14

Using specific phospholipid transporters flippases and floppases

Requires energy (ATP)

Question 15

Which direction to flippases and floppases move phospholipids

Answer 15

Flippase moves PS from outer to inner

Floppase moves PS from inner to outer

Question 16

Why doesn’t turning off flippases/floppases result in enough PS in the outer leaflet to activate prothrombinase

Answer 16

It would be too slow and you need rapid movement of PS to the outer leaflet or you would bleed to death

Question 17

How is phospholipid asymmetry destroyed to have enough PS on the outer leaflet

Answer 17

Destroyed enzymatically using phospholipid scramblase

Question 18

How is phospholipid scramblase activated

Answer 18

Normally inactive, and is activated by increased calcium which occurs in response to damaged blood vessels

Question 19

What is Scott Syndrome? Where is the probelm

Answer 19

Inherited blood coagulation disorder

Extremely rare where the conversion of prothrombin to thrombin is defective (the coagulation factors are intact)

Defect in the membrane lipid scrambling in activated platelets so there is not enough PS in the outer leaflet

Defect in TMEM16F

Question 20

How do you measure PS exposure

Answer 20

Annexin V labeling

Question 21

What is Annexin V labeling

Answer 21

Microscopy/visualization method

Annexin V bind to PS in the presence of calcium

Annexin V stagged with a fluorophore

Question 22

What is scramblase activity like in Scott Syndrome

Answer 22

Scramblase activity is normal

Question 23

What is TMEM16F? How does a defective TMEM16F affect Scott Syndrome

Answer 23

It is an ion channel to transport [Ca] into the cell

When the coagulation factor is activated, TMEM16F causes increased intracellular calcium

Increased calcium is required for the calcium-dependent activation of PL scramblase

Question 24

What is the mutation in TMEM16F in scott syndrome

Answer 24

G –> T nonsense mutation in the TMEM16F gene

Makes a severely truncated protein which is non-functional

Question 25

How can PS exposure be used as a tool for cancer therapy

Answer 25

Cancer cells have more PS present in the outer leaflet

This increased PS exposure could be used to visualize cancer cells by adding SapC-DOPS which binds to surfaces rich in PS

Question 26

Why can’t cancer cells maintain phospholipid asymmetry?

Answer 26

They cannot maintain asymmetry because they have low flippase activity and higher [Ca] which increases scramblase activity

Question 27

What is Sap C-DOPS

Answer 27

Artificial complexes of lipids and proteins that contains saposin C and dioleoyl-PS

It binds to membranes containing phosphatidylserine

Question 28

How can SapC-DOPS be used

Answer 28

can be used to kill cancer cells and does not affect healthy cells

SapC is internalized by cells and activates cell death pathways

No serious adverse effects