Protein C

Question 1

Why is the protein C pathway important?

Answer 1

The TFPI pathway can only inhibit small amounts of coagulation and once there is sufficient thrombin to overcome

Question 2

What is the APC?

Answer 2

Activated protein c pathway

Question 3

Which molecules are involved in the APC pathway?

Answer 3

Protein C
Thrombin
Thrombomodulin
FVa and FVIIIa

Question 4

Which molecules does APC target?

Answer 4

The tenase complex

Question 5

How does the APC pathway work?

Answer 5

Thrombin is exposed due to vessel injury, on the endothelial cells it binds to thrombomodulin - thrombin becomes anticoagulant in mechanism
Protein C binds to EPCR with high affinity
Protein C binds to TM and thrombin cleaevs protein C to APC and the activation peptide is removed
APC is released and another protein C binds the EPCR
Once released it binds protein S which is a cofactor - it can now inactivate 5a and 8a

Question 6

What is the importance of the Protein C pathway?

Answer 6

Reduces and regulates the spread of the haemostatic plug

Question 7

What is protein C?

Answer 7

It is a vitamin K-dependent serpin zymogen

Question 8

What are the structural features of protein C?

Answer 8

Conserved domain structure with FVII FX and FIX

Made up of a gla domain but due to vit k dependence is not able to bind phospholipids in the absences of vit K

Question 9

What are the structural features of protein S?

Answer 9

Sex hormone binding globulin domain
4 EGF domains
thrombin sensitive region
gla domain - this is essential to its function

Question 10

What effect does binding of thrombin to thrombomodulin have?

Answer 10

The binding leads to inactivation of thrombomodulin and activation of protein C

Question 11

What is the importance of chondroitin sulphate domain on TM for activation of protein c?

Answer 11

It allows the thrombin to bind to thrombomodulin with hug affinity - it has additional exosites

Question 12

What is the mechanism of thrombomodulin in the activation of protein C?

Answer 12

It aligns the active site of thrombin to allow protein c activation

Question 13

Where is thrombomodulin found?

Answer 13

It is concentrated on the surface of endothelial cells and is adjacent to the site of injury

Question 14

Which are the EGF domains essential for function of TM?

Answer 14

EGF 456

4 binds the protein C serine protease domain

Question 15

What is the cleavage site of protein C which leads to its activation? What molecule cleaves protein C?

Answer 15

It cleaves the activation peptide at specific Arg169 - this activates protein c to APC, it is cleaved by thrombin

Question 16

Why does cleaving lead to dissociation of APC from EPCR?

Answer 16

Cleavage of the activation peptide leads to a decrease in the affinity of the binding which leads to APC being released from EPCR

Question 17

How does APC inactivate FVa?

Answer 17

It cleaves at specific Arg residues (306/506/679)

Question 18

Where does inactivation occur?

Answer 18

On anionic phospholipid surfces - activated platelet and endothelial cell membranes at the edge of the injured area

Question 19

Explain the biphasic nature of inactivation of FV

Answer 19

Arg 506 is rapid cleavage and reduces activity of cofactor by 40%
Arg 306 cleavage is slow and this abolishes FVa activity

Question 20

How does protein S act as a cofactor?

Answer 20

It has a high affinity for phospholipids so increases the affinity of APC to the phospholipid surface which localises it to the site of FV and FVIII

It reorientates the active site to increase slow cleavage of FVa at 306 thus accelerating its inactivation, overall x20 increase of inactivation

Question 21

How does APC inactivate FVa?

Answer 21

It cleaves at specific Arg residues (306/506/679) in the A2 domain

Question 22

How is FVa protected from inactivation by APC?

Answer 22

It is bound to Xa in circulation wich protects it

Question 23

How important is protein S to function of APC?

Answer 23

APC activity is basically entirely dependent on protein S

Question 24

How does APC inactivate FVIIIa?

Answer 24

Cleavage at Arg336 and 562 but this is relatively inefficient

Question 25

How is APC inactivated?

Answer 25

It has a half-life of 15-30 mins this is relatively slow

the inhibitors are alpha1 antitrypsin, alpha 2 macroglobulin and PCI

Question 26

What is FV Leiden?

Answer 26

It is a mutaion associated with APC resistance, there is a change in one of the cleavage inactivation sites - 506, therefore the fast stage of inactivation is lost

Question 27

Describe the structure of thrombomodulin

Answer 27

Lectin like domain
6 EGF
chondroitin/ser/threo region
transmembrane region