Apoptosis Control Proteins (L5)

Question 1

How is apoptosis activated extrinsically?

Answer 1

Killer lymphocyte carrying Fas ligand binds and activates Fas proteins on surface of target cell. Adaptor proteins bind to intracellular region of Fas proteins, causing aggregation of procaspase-8 molecules. These then cleave on another to initiate caspase cascade.

Question 2

How is apoptosis activated intrinsically?

Answer 2

Mitochondria release cytochrome c, which binds to and causes aggregation of adaptor protein (Apaf-1). Apaf-1 binds and aggregates procaspase-9 molecules, which leads to cleavage of these molecules and the triggering of caspase cascade

Question 3

Name some of the members of the BCL-2 family and their function

Answer 3

Bcl-2 and bcl-Xl- anti-apoptotic

Bax, Bak, Bid, Bim- pro-apoptotic

Question 4

What is the structure of BCL-Xl?

Answer 4

Seven alpha helices arranged in three layers
Two central, predominantly hydrophobic helices surrounded by amphipathic helices
BH1-BH3 regions clustered on one side of molecule and form hydrophobic patch

Question 5

What are the features of the BCL-Xl/Bak peptide complex?

Answer 5

Bak BH3 peptide adopts an alpha-helix that fits into a hydrophobic cleft formed by residues in the BH1,2 and 3 regions of BCL-Xl
Hydrophobic and electrostatic interactions

Question 6

What are some of the general roles of caspases?

Answer 6

Responsible for most of the changes seen in apoptosis

Can function as initiators of death cascade or as effectors

Question 7

Describe the structure of a procaspase

Answer 7

N-terminal pro-domain, large subunit (p20) and small subunit (p10) domains

Question 8

How are caspases usually activated?

Answer 8

Proteolytic cleavage of inactive procaspase between p20 and p10 domain, and usually also between prodomain and p20

Question 9

What is the simplest way to activate a procaspase?

Answer 9

Expose it to another previously activated caspase molecule

Question 10

What are the three distinct specificity groups?

Answer 10

WEHD (1,4,5,13)
DEXD (2,3,7)
(I/V/L)EXD (6,8,9,10)

Question 11

What is the structure of caspase-3 dimer?

Answer 11

Caspase fold consists of a large subunit (p17) and a small subunit (p12)
Each p17/12 heterodimer comprises 6 beta-strands, that form a twisted beta-sheet structure, with 2 alpha helices on one side and 3 alpha-helices on the other

Question 12

What is the structure of caspase-8 tetramer?

Answer 12

Similar to caspase-3 and consists of a (p12/p18)2 tetramer composed of 2 heterodimers
C terminus os large subunit is far away from N terminus of small subunit in the heterodimer but is very close to the N terminus of the small subunit in the adjacent protein

Question 13

Where is the recognition site for the substrates of caspases?

Answer 13

Predominantly in cleft formed by loop regions of the p18 and p12 subunits

Question 14

What is the catalytic mechanism of substrate recognition within caspases?

Answer 14

Activation of the active site cysteine (cys285 in caspase-8), which is polarised by a histidine residue (His237)

Question 15

Describe the substrate specific pocket for caspase-1

Answer 15

Large hydrophobic pocket that can accommodate large hydrophobic residues

Question 16

Describe the substrate specific pocket for caspase-3

Answer 16

Relatively narrow pocket that forms H bonds with the Asp preffered at the P4 position

Question 17

Describe the substrate specific pocket for caspase-8

Answer 17

Specificity pockets for P3 and P4, resulting in a preference for Glu at P3 and small hydrophobic amino acids at P4