IT - Proteases II

Question 1

What is the function of the proteasome?

Answer 1

Ubiquitin-dependent degradation of proteins

• Requires ATP

Question 2

How was the structure of the proteasome first determined?

Answer 2

Single-particle analysis in electron microscopy

Question 3

What are the two main components of the proteasome cap, and what are their functions?

Answer 3

• Rpn: senses ubiquitination state
• Rpt: ATPase

Question 4

What is the function of the central core of the proteasome?

Answer 4

It houses the active protease with a large cavity open on both sides.

Question 5

In the eukaryotic proteasome central core, which subunits are active?

Answer 5

Only b1, b2, and b5

Question 6

What type of enzyme is the proteasome, and what is unusual about its catalytic triad?

Answer 6

It’s a triad enzyme, but the nucleophile is a threonine (not the usual serine)

Question 7

Whats involved in Peptidoglycan synthesis? (2)

Answer 7

Synthesis I

• Peptidoglycan chain 1 attacked by Serine Transpeptidase and D-Ala removed

Synthesis II

• Peptidoglycan chain 2 attacks carbonyl on 1 and Serine Transpeptidase is removed

Question 8

How does penicillin specifically target the transpeptidase?

Answer 8

The reactive beta-lactam ring forms a covalent bond with the enzyme, irreversibly inhibiting it

Question 9

Function of b-lactamases (2)

Answer 9

• b-lactamase cleaves the b-lactam of penicillin, leading to antibiotic resistance
• b-lactamase, like transpeptidase, is a triad enzyme

Question 10

How do some antibiotics overcome B-lactamase resistance?

Answer 10

They target and inactivate the B-lactamase enzyme itself (e.g., clavulanic acid)

Question 11

How does the similarity of the triad mechanism allow for the development of specific enzyme inhibitors?

Answer 11

Despite similar mechanisms, different enzymes have varying substrate specificities and fine-tuning

• This allows targeting specific enzymes with inhibitors that fit their active site shape

Question 12

What does the existence of similar catalytic triads in unrelated pathways suggest about evolution?

Answer 12

It suggests convergent evolution, where the same efficient solution (triad mechanism) has evolved independently multiple times

Question 13

Describe the steps involved in the aspartate protease mechanism

Answer 13

1. General base polarizes a water molecule
2. Nucleophile attacks the carbonyl carbon of the peptide bond
3. A tetrahedral intermediate forms
4. Leaving group is protonated
5. Tetrahedral intermediate collapse
6. Products leave the enzyme.

Question 14

Give some examples of aspartate proteases (3)

Answer 14

• Renin
• Pepsin
• HIV-1 Protease

Question 15

How do HIV protease inhibitors work?

Answer 15

They mimic the structure of the tetrahedral intermediate, blocking the active site and preventing HIV from processing viral proteins

Question 16

How can HIV develop resistance to protease inhibitors?

Answer 16

Mutations in the HIV protease gene can alter the active site shape, rendering the inhibitor unable to bind effectively

• Mutation of Val48 confers 13.5-fold resistance
• Mutation of Met90 confers 3-fold resistance

Question 17

Briefly explain Lewis acids and bases

Answer 17

Lewis bases donate electron pairs

Lewis acids can react with Lewis bases to form Lewis adducts

Question 18

Give some examples of metalloproteases and their functions

Answer 18

• Thermolysin (bacterial protease),
• **Matrix metalloproteases **(MMPs, involved in signaling and wound healing)

Question 19

Where is the APP (Alzheimer Precursor Protein) located?

Answer 19

In synapses

Question 20

What are the two main pathways for APP processing, and how does one contribute to Alzheimer’s disease?

Answer 20

• Alpha-secretase pathway (non-amyloidogenic)
• Beta-secretase pathway (contributes to Alzheimer’s disease)