WEEK 4 - Beta-lactamase Inhibitors Flashcards
describe the chemical properties of a drug, which may include its bonding, shape, functional groups, stereochemistry, ionisation (pKa) and lipophilicity (LogP/D)
RECAP: Bacterial cell targets (for drug development)
YEAR 2 Lecture Notes
Can develop
1. Cell wall synthesis inhibitors
- cell wall has peptidoglycan mesh (glycan strains cross linked together by PBP)
- mesh proveds rigidity
- penicillin was 1st antibitoitc discovered to weaken mesh = cell lysis = death
2. Protein synthesis (ribosomes) inhibitors
3. DNA synthesis inihbitor
4. Cell membrane disruptor
Describe bacterial cell wall composition: Peptidoglycan
inc. process PBP works
MoA of Bacterium:
- Bacterial cell wall uses PBP (penicillin-binding protein) to bind / cross link glycan strains (attached to NAM)
- short peptide chains form between D-Ala and DAP on each glycan strains = peptidoglycan fomed
- Peptidoglycan / crossliniking prevents cell from bursting under pressure = bacteria able to survive
- keeps cell wall rigid
PBP: acylation-deacylation process of PBP
- Serine from PBP attacks peptide bond = SP3 tetrahedral intermediate formed
= acylated form of enzyme is formed with -ive charge on oxygen
- Oxyanion hole is a pocket in active site of enzyme that stabilises -ive charge on oxygen
- Negative charge accelerates enzyme reaction / catalyses= deacylation of enzyme is enabled = stable form of enzyme is formed
- Crosslinking can occur
NOTE:
- PBP is an enzyme transpeptidase
- PBP is membrane bound
How do B-lactams work
(Antibitoic)
Prevent deacylation occuring = NO crosslinking will occur
- Serine on PBP attacks beta-lactam ring (instead of peptide / glycan strains)
- b-lactam acylates PBP (enzyme) = -ive tetrahedral intermediate is formed
Preventing deacylation causes:
- Inactivation of PBP as it gets stuck in acylated form (tetrahedral form)
- Deacylation of enzyme can NOT occur = crosslinking between glycan strains does NOT happen
- Bacterial cell wall is weakened = lysis from osmotic pressure = cell death
What are b-lactamases
Enzymes that hydrolyse the beta lactam ring (in antibiotics) AND releases the hydrolysed form of drug
- Hydrolysation prevents PBP from binding to antibitoic
- inhibiting the antibiotic + making it inactive
- Lactamases are proteins found in the space beween the inner + outer walls of bacteria
- NOT membrane bound
Beta lactamases are classified into 2 types:
1. SBLs (serine b-lactamases)
- Class A, C and D
- have a deep active site
- have serine (+ lysine) active site
2. MBLs (metallo b-lactamases)
- Class B (3 subtypes: B1, B2, B3)
- have a shallow active site
- have 2 zinc at active site + hydroxyl (OH-)
How do beta-lactamases promote deacylation of PBP
Aceylation-deacylation process
1. Serine on PBP attacks
2. Tetrahedral intermediate is formed
3. Lactamases can proceed further (PBP in prescence of b-lactam can NOT get past step 2)
4. Lactamase re-arranges its conformation + attacks again knocking off the b-lactam from the serine
5. Deacylation occurs = cross linking occurs
How to combat b-lactamase
- Develop b-lactamase resistant compounds
2.Develop b-lactamase inhibitors
Develop b-lactamase resistant compounds
Combatting b-lactamase, 3 solutions
1st solution:
- Developing penicillins with large side chains
- prevents lactamase binding BUT still allows PBP to bind
NOTE:
- resistance developed, mutated PBP emerged = MRSA
- mutated bacterium were resistant to b-lactamase resistant penicillins
2nd solution:
- Developed cephalosporins
NOTE:
- resistance developed, ESBLs emerged
3rd solution:
- Developed carbapenems
- They binded + inactivated PBP and b-lactamases (inc. ESBLs)
- Used widely as last resort
Develop b-lactamase inhibitors
2 Examples
They bind to b-lactamse, inactivating it
- Clavulanic Acid
- Binds well to b-lactamase (but not PBP)
- covalently binds
- serine from lactamase attacks beta lactam ring on c.acid, opening ring
- another serine attacks c.acid
- c.acid cross links the 2 serine = deatcivated lactamase
- Used in combo. with b-lactam antibiotics (that were susceptible to b-lactamase)
- Active against gram +ive and -ive Class A SBLs - Sulbactams
- Same MoA as C.acid (acylates initial b-lactamase + reacts further with another serine from lactamase)
- Used in combination
- Active agaisnt Class A SBLs, ESBLs
- Inactive against gram -ive
What caused clavulanic acid resistance
B-lactamases had mutations at active site = lost binding ability to c.acid
- due to loss of salt bridges + ionic interactions
- e.g. TEM-1 lactamase mutation of Argenine to Serine
Carbapenemase B-lactamases
Inc. 3 classes of lactamase, how they work
- Overuse of carbapenems led to resistance and CPEs (carbapenemases)
- Resistance occured in multi-drug resistant gram -ive bacteria
What are the 3 classes of carbapenemases:
1. Class D - SBLs
2. Class B1 - MBLs
3. Class A - SBLs
How SBL carbapenemase work
- Rotate 6a-hydroxyethyl group away from glutamate and water
= reaction can occur
- Rigid active site using disulfide bonds to hold C=O group in oxyanion hole = deacylation can occur
DBOs (diazabicyclooctanones) INFO
- MoA unclear
- negative charged molecule that fits into lactamase pocket - Inhibit most Class A, B and D b-lactamases
- Some have PBP activity
Examples:
- Avibactam
- Relebactam
NOTE:
- DPO resistance has been reported
Cyclic Boronates INFO
- Inhibit lactamases
- forms covalent bonds with serine
- not active against MBL
STRUCTURE
- form 3 SP2 hybridised orbitals
- allows inital binding
- When reacts with nucleophile = SP3 (tetrahedral) is formed
- tetrahedral form is stable to hydrolyse
- mimics acylation intermediate (-ive charge)
- Form flat triganol structure
Example:
- Varborbactam
Issue with inhibiting MBLs
MBLs = metallo b-lactamases
NO clinically approved drugs
PROBLEMS with INIHBITING MBLs:
- Have a shallow active site
- Broad similiarity to other proteins
- No amino acids directly involved in catalysis = can NOT target specific one
Why are MBLs a PROBLEM :
- Rise of potent undruggable B1 carbapenemases
- NDM (new delhi MBL)
- rapid spread, epidemic in india
- can hydrolyse all b-lactams
What are the 3 strategies to target MBLs
- Displace hydroxyl (OH-) nucleophile
- Extract / remove zinc ions
- Design small molecules to block active site and inhibit MBLs
