L12 - enzyme as drug targets Flashcards
What do simple models of reversible enzyme inhibition assume?
- 1 inhibitor molecule binds to enzyme
- inhibitor binds is reversible
- linear kinetics observed
- inhibitors are dead-end
What do inhibitors resemble?
substrate in structure
what do inhibitors affect
either Km, Vmax or both
what is potency measured by?
Ki value (conc of inhibitor that causes a 2 fold change in Km or Vmax
what is Ki values related to?
binding energy (deltaG)
what do smaller Ki numbers mean?
tighter bonding (higher potency)
what do many potent inhibitors resemble?
the transition state of reaction
what does the development of new inhibitors require?
understanding of enzyme mechanism
what is an example of of an inhibitor?
proline racemase
- transition state is relatively flat
- inhibition by pyrrole-2-carboxylate
what are the forms of inhibition?
- competitive
- non-competitive
- mixed competitive
- uncompetitive
what is the most common form of inhibition?
competitive inhibition
what are the plots like for competitive inhibition?
- Km increases (x int changes)
- Km (I) = Km (1 + [I]/Ki)
- no change in Vmax (y int the same)
when does inhibition decreases in competitive inhibition?
with increased substrate concentration
what is non-competitive inhibition uncommon for?
single substrate enzymes
What are plots like for non-competitive inhibition?
- Vmax is reduced
- Vmax/Km (I) = Vmax/Km(1 + [I]/Ki)
- Km not affected
When is inhibition reduced in non-competitive inhibition?
it’s not reduced by increasing substrate
What is a common form of inhibition?
mixed competitive inhibition
what are plots like for mixed competitive inhibition?
- Km increased (comp)
- Vmax decreased (non-comp)
what does the alpha factor define?
the competitive and non-competitive components
what is an uncommon mode of inhibition?
uncompetitive inhibition
How does the uncompetitive inhibition?
inhibitor binds to the ES complex
What are plots like for uncompetitive inhibition?
- parallel lines in lineweaver-burk plot
- both Km and Vmax reduced by same amount
What happens when there’s an increase in substrate concentration in uncompetitive inhibition?
it increases inhibition
what do dose response curves measure?
drug potency at a fixed substrate concentration
what is IC50?
conc of drug required for 50% reduction in activity
- not the same as Ki
what does the value for IC50 depend on?
assay conditions
- especially substrate conc
when does tight binding inhibition occur? and what does this mean?
when the conc of active enzyme and inhibitor are similar
- conc of free inhibitor doesn’t approximate to total inhibitor
When does the Ki value increase?
with increasing enzyme conc
what does tight-binding inhibition often have? and what does this mean?
- very slow onset
- the enzyme inhibitor complex can have a very long half life
- infrequent dosing regiments can be used
what are examples of irreversible inhibitors?
penicillins and other B-lactam antibiotics
what are enzymes inhibited by with penicillins?
penicillin-binding proteins (PBPs)
what do irreversible inhibitors act as?
pseudo-substrates but get “stuck”
- at acyl-enzymes intermediate)
what is kinetic behaviour like with irreversible inhibitors?
more similar to chemical systems
