Lecture 13 - Enzyme inhibitors, therapeutics and diagnostics Flashcards
1) What is Enzyme Competitive Inhibition?
2) What does the effect on rxn rate depend on?
3) What happens at zero order kinetics?
4) What’s the effect on Vmax as a result of inhibitor?
1) substrate and inhibitor compete for binding at the substrate binding site
2) this effect depends on concentrations of substrate and inhibitor and the relative affinities of S and I for that site
3) where substrate and enzyme are balanced, if add a little inhibitor, won’t matter, rxn will still reach the Vmax
4) nothing, Vmax is unchanged
True or False:
Rxns regarding competitive inhibitors are reversible.
True
Describe Ethylene Glycol toxicity
1) lots of EG in MN
2) most common in cats and dogs
3) most intoxications come from ingestion of antifreeze (95% EG)
4) Only requires small amount ingested to be lethal/toxic
5) EG is rapidly absorbed in GI tract, however in dogs peak blood concentrations of it take about 3 hrs after ingestion
1) Why is it good that it takes about 3 hrs in dogs to reach peak high blood concentrations of EG?
2) what makes it glow?
- how is that helpful?
1) 3 hrs is relatively long, lots of EG is excreted out during that time and avoid being digesteed and absorbed
2) there are fluorescent properties within EG –> sometimes can see this represented in urine
Explain EG degradation pathway
EG –> (alcohol dehydrogenase) –> glycoaldehyde (SLOW)
Glycoaldehyde –> glycolic acid (RAPID)
Glycolic acid –> glyoxylic acid (SLOW)
Glyoxylic acid –> formic acid, CO2, glycine, serine and OXALATE (RAPID)
1) What are the rate-limiting steps of the EG pathway?
2) why is it good these steps occur within first few hrs
3) Which products are most toxic/harmful?
4) What happens with the FIRST rate-limiting step?
5) What results from the formation of oxalate?
1) EG –> glycoaldehyde
glycolic acid –> glyoxylic acid
2) they are slower steps in the pathway and so allow time for lots of EG to be excreted instead of being processed
3) glycollic acid and oxalate
4) since it is a slower step, Glycolic acid accumulates and leads to acidosis –> kidney damage
5) oxalate crystallizes with Ca+2 in kidney –> damaging to kidney, heart, lungs and brain.
can look in urine sometimes and see crystals
1) What compound is used to prevent metabolism of EG?
2) How does it work?
3) What species is it effective in?
4) What species is it ineffective in and what do they use in place?
1) 4-MP (4-methylpyrazole)
2) 4-MP acts as a competitive inhibitor for alcohol dehydrogenase, so competes with EG to bind with alcohol dehydrogenase –> activity of that enzyme decreases –> prevents metabolism of EG
3) effective in dogs without the side effects of ethanol
4) 4-MP is inefffective in cats at canine dosage. Ethanol is treatment of choice for cats (acts as competitive inhibitor of alcohol hydrogenase) but still comes with same side effects.
Research has found that doctors can use 4-MP but need 6x amount used in dogs and also need to start with an additional initial dose –> safe and effective in cats at this dose
1) What is enzyme uncompetitive inhibition?
2) What is the effect of high concentrations of S on I?
3) can you reach Vmax?
1) Inhibitor binds at a distant site when S is on substrate, therefore can’t form the product (binds to transition complex (ES)
2) nothing, S and I aren’t competing so high S doesn’t affect I or reverse I’s effect.
3) No, no matter how much S you add, you won’t reach Vmax
1) What are irreversible inhibitors?
2) give an example
1) they combine with or destroy a fn gp on an enzyme. They commonly form a covalent bond btw enzyme and the inhibitor
2) DFP (involved with serine proteases) irreversibly inhibits AchE (acetylcholinesterase) by binding and destroying it
1) What’s a special class of irreversible inhibitors?
2) What do they do?
1) Suicide inhibitors
2) they mimick transitional state or look like prdt –> they carry out first few chemical steps of normal enzymatic rxn then they get locked in to enzyme –> TS can’t transform to prdt but instead inhibitor binds irreversibly
1) What is the goal of designing a selective inhibitor?
2) What are the two cyclooxygenase enzymes and what do they do?
3) whhat is drug companies goals for these two?
1) design a drug that resembles the natural substrate and can fit specifically into substrate binding site on key regulatory enzymes and inhibit rxns.
can also design uncompetitive inhibitor
2) COX-1 (good) and COX-2 (bad–>arthritis) generate prostaglandin metabolites which are good and bad inflammatory intermediates (PGI2 - prevents clotting and thromboxane - clotting)
3) selectively inhibit COX-2 to decrease levels of bad prostaglandins without side effects (VIOXX)
aspirin - irreversible inhibitor
1) What are Toxins/poisons?
2) what are some common examples?
3) How do OPs work?
4) What are the effects of OP poisoning?
1) they can be inhibitors of key metabolic and physiological processes –> selective action on specific proteins and enzymes
2) Insecticides (organophosphates or OPs) - irreversible AChE inhibitors
3) AChE degrades neurotransmitter acetylcholine –> inhibit AChE –> build up of acetylcholine in synaptic cleft –> neuromuscular paralysis
Once OPs are bound they can’t break down
4) SLUDGEM = salivation, lacrimation, urination, defecation, gastrointestinal motility, emesis (vomiting), and miosis (pupils contract)
1) What’s an antidote for OP poisoning?
2) True or False: Atropine binds to AChE?
3) What is atropine’s effect on AChE?
4) True or False: Atropine is the treatment of choice for OP poisoning.
1) Atropine
2) False - atropine binds to post-synaptic acetylcholine RECEPTORS and prevents accumulated acetylcholine from binding to receptor –> suppresses the effect of excess acetylcholine (blocks signal)
3) nothing –> AChE is irreversibly inhibited by OP
4) False - atropine is not used a lot. even though it works well, it has a lot of side effects and the drug is short-lived. typically use muscle relaxors and supportive care
1) Describe allosteric sites.
2) What are the two types of allosteric enzymes?
3) What happens to enzyme + substrate?
1) unique region of enzyme that is different from the substrate-binding site
2) Allosteric effector and inhibitor
3) allosteric enzyme causes a conformational change in E which results in an increased affinity for S (effector) or decreased affinity for S (inhibited)
***usually the substrate is being modified NOT allosteric enzyme
What are most allosteric enzyme systems found as?
What does this mean?
commonly are dimers or trimers (multiple subunits)
when allosteric enzyme binds to first subunit, it increases/decrease the affinity of both subunits
1) What are the two models for Allosteric enzymes?
2) Describe them
1) Concerted model and sequential model
2) Concerted model –> there is a ligand bound and unbound state and both pre-exist at the same time (simultaneously)
both states are in equilibrium with each other and enzyme can flip back btw states
once bound, the rxn shifts to right toward ligand-bound state
Sequential model - only taut (unbound form pre-exists) –> all in unbound state
1 substrate bound to subunit affects next subunit (hemoglobin and positive cooperativity)
1) where are allosteric enzymes found?
2) How can we use Allosteric enzymes in clinical setting?
1) sequestered in mitochondria, found in blood vessels
2) enzyme assays –> look at blood chemistry –> see where creatinine kinase is released and see where LDH and HBDH are released later –> indicate why someone had heart attack
