2. Enzyme kinetics Flashcards
Pos cooperativity vs neg cooperativity. Know their graphs and examples of pos and noncooperativity
inc affinity for subsequent S, Hill’s coeff (n) > 1; ex: hemoglobin vs dec affinity for subsequent S, Hill’s coeff (n) < 1. Hill’s coeff (n) = 1 means enzyme has no cooperative binding; ex: myoglobin
How to find K_a vs K_d?
[PL]/[P]*[L] vs 1/K_a
Myoglobin vs hemoglobin. Know the O2 binding curves
Stores oxygen, like enzyme saturation curve (hyperbolic) vs transports oxygen, sigmoidal, tense state - rigid, hard to bind O2 & relaxed state - easy to bind to O2 —> pos cooperativity; pO2 = x axis, sat O2 = y axis
Know high affinity vs low affinity on graph. What happens when you dec pH?
High = towards left top vs low = towards right bottom. Lower pH –> more p+ –> more oxygen released –> lower affinity for oxygen –> higher K_d (graph shifts to right)
How does Keq and deltaG relate to substrates and products?
Small Keq –> substrates = favored –> positive deltaG
Large Keq –> products = favored –> negative deltaG
What’s the general idea of Michaelis and Menten? What’s the eqn? What are the 4 asmptns?
find K_M at 1/2Vmax. V = (Vmax*[S])/(Km+[S]) where V=initial vel
1) this eqn = only used to describe initial rxn vel
2) steady state approximation (ie. [S]»>[P])
3) free ligand approximation (ie. [S]»>[E])
4) rapid equil approximation (ie. Lower Km means higher enzyme-substrate affinity)
Competitive vs uncompetitive vs noncomp vs mixed inhibitors. Know what they look like graphically (ie. LINEWEAVER BURK PLOTS)
competes w/ S, inc Km, (inc S does overcome inhibitor) vs binds to allosteric site of ES, dec Vmax and Km (inc S doesn’t overcome inhibitor) vs either binds to allosteric site of E or ES, dec Vmax, (inc S doesn’t overcome inhibitor) vs either binds to allosteric site of E or ES, inc/dec Km and dec Vmax
Know lock and key vs induced fit model
specific shape for S to bind to active site of E vs E changes shape to accommodate S to bind; E changing shape = endergonic since it requires energy, E going back to nml shape = exergonic
Cofactors or coenzymes vs apoenzymes vs holoenzymes
proteins that bind to enzymes to do their respective thing vs enzymes that need the cofactors/coenzymes to do their thing vs cofactors/coenzymes + apoenzymes
What factors affect enzyme activity?
High temp (optimal = 37 degrees C/310 K), pH not 7.4 physiologically and affecting ionization, high salinity affecting H bonds and ionic bonds
What does Km tell you?
[S] at half Vmax
Know the enzyme saturation graph. Know what comp and noncomp inhibition looks like on nml graph
x axis = [S], y axis = rxn rate
Noncomp is lower but same shape, comp has LESS steeper slope but same Vmax
Lyases vs ligases vs hydrolases vs transferases vs dehydratases vs isomerases vs restriction enzyme/endonuclease aka restrictase vs hydroxylase vs decarboxylase vs phosphorylase vs kinase
cleave w/o water or e- transfer vs linking molec together vs cleaves w/ water vs transfers functional group from one molec to another vs removes water vs inter conversion of isomers vs cutting DNA into fragments at restriction sites vs facilitates hydroxylation vs removes carboxyl group and release CO2 vs add inorganic phosphate to another molec vs transfers phosphate group to another molec typically from ATP
T/F: hemoglobin has lower affinity to O2 than myoglobin
T
What is irreversible inhibition?
When active site = unavailable for prolonged time —> enzyme permanently lost function —> need newly made enzymes to do the job
What are allosteric enzymes? Allosteric activator vs allosteric inhibitor?
Enzymes with active site + multiple allosteric sites. Causes shift in more available active site vs causes shift in less available active site; both of these = allosteric regulation
How does ideal temp of a rxn change w/ or w/o a catalyst?
Ideal temp = lower for rxn w/ catalyst than without. Higher temp is ideal for rxn without catalyst bc higher kinetic energy can overcome activation energy. But with the catalyst, you can’t have too high of a temp or else you’ll denature catalyst
As hemoglobin unloads oxygen, what happens to affinity of remaining oxygen?
Lowers affinity
Enzymes stabilizes transition state —> lowers activation energy —> faster rxn rate
Enzymes can affect kinetics BUT NOT THERMODYNAMICS (ie. They don’t affect deltaG)
A single enzyme doesn’t catalyze a variety of rxns cuz they’re usually highly specific
Factors that affect Vmax. How does [S] affect enzyme’s rxn rate?
Only enzyme and enzyme conc. Inhibitors could dec V, but doesn’t necessarily change Vmax
higher [S] inc rxn rate by enzyme until all enzymes = saturated (by that time, only [E] can determine rxn rate)
What are some ways to turn a tense enzyme (ie. Off) into a relaxed enzyme (ie. On)?
Phosphorylation of enzyme, allosteric regulation (allosteric activator/inhibitor)
What’s a substrate analog?
A molec resembling to nml substrate that binds to enzyme’s active site —> competitive inhibition
If there are equal parts/conc of substrate and inhibitor, how do you know which one the enzyme will take?
Pick the one that will stabilize transition state (ie. ES complex) better
Feedforward stimulation vs feedback stimulation vs feedback inhibition
Early intermediate triggers enzyme to inc activity vs later intermediate triggers enzyme to inc activity vs intermediate triggers previous intermediate to dec activity
Bohr effect
Hemoglobin’s affinity to O2 based on pH and CO2; its affinity to O2 dec when low pH and high CO2
Eqn for catalytic efficiency
Kcat/Km
How to shift right on oxyhemoglobin dissociation curve?
Exercise is the right thing to do: increase [H+], increase PaCO2, increase temp, increase 2,3 bisphosphoglycerate
