2. Enzyme kinetics

Question 1

Pos cooperativity vs neg cooperativity. Know their graphs and examples of pos and noncooperativity

Answer 1

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

Question 2

How to find K_a vs K_d?

Answer 2

[PL]/[P]*[L] vs 1/K_a

Question 3

Myoglobin vs hemoglobin. Know the O2 binding curves

Answer 3

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

Question 4

Know high affinity vs low affinity on graph. What happens when you dec pH?

Answer 4

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)

Question 5

How does Keq and deltaG relate to substrates and products?

Answer 5

Small Keq –> substrates = favored –> positive deltaG

Large Keq –> products = favored –> negative deltaG

Question 6

What’s the general idea of Michaelis and Menten? What’s the eqn? What are the 4 asmptns?

Answer 6

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)

Question 7

Competitive vs uncompetitive vs noncomp vs mixed inhibitors. Know what they look like graphically (ie. LINEWEAVER BURK PLOTS)

Answer 7

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

Question 8

Know lock and key vs induced fit model

Answer 8

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

Question 9

Cofactors or coenzymes vs apoenzymes vs holoenzymes

Answer 9

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

Question 10

What factors affect enzyme activity?

Answer 10

High temp (optimal = 37 degrees C/310 K), pH not 7.4 physiologically and affecting ionization, high salinity affecting H bonds and ionic bonds

Question 11

What does Km tell you?

Answer 11

[S] at half Vmax

Question 12

Know the enzyme saturation graph. Know what comp and noncomp inhibition looks like on nml graph

Answer 12

x axis = [S], y axis = rxn rate

Noncomp is lower but same shape, comp has LESS steeper slope but same Vmax

Question 13

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

Answer 13

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

Question 14

T/F: hemoglobin has lower affinity to O2 than myoglobin

Answer 14

T

Question 15

What is irreversible inhibition?

Answer 15

When active site = unavailable for prolonged time —> enzyme permanently lost function —> need newly made enzymes to do the job

Question 16

What are allosteric enzymes? Allosteric activator vs allosteric inhibitor?

Answer 16

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

Question 17

How does ideal temp of a rxn change w/ or w/o a catalyst?

Answer 17

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

Question 18

As hemoglobin unloads oxygen, what happens to affinity of remaining oxygen?

Answer 18

Lowers affinity

Question 19

Enzymes stabilizes transition state —> lowers activation energy —> faster rxn rate

Answer 19

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

Question 20

Factors that affect Vmax. How does [S] affect enzyme’s rxn rate?

Answer 20

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)

Question 21

What are some ways to turn a tense enzyme (ie. Off) into a relaxed enzyme (ie. On)?

Answer 21

Phosphorylation of enzyme, allosteric regulation (allosteric activator/inhibitor)

Question 22

What’s a substrate analog?

Answer 22

A molec resembling to nml substrate that binds to enzyme’s active site —> competitive inhibition

Question 23

If there are equal parts/conc of substrate and inhibitor, how do you know which one the enzyme will take?

Answer 23

Pick the one that will stabilize transition state (ie. ES complex) better

Question 24

Feedforward stimulation vs feedback stimulation vs feedback inhibition

Answer 24

Early intermediate triggers enzyme to inc activity vs later intermediate triggers enzyme to inc activity vs intermediate triggers previous intermediate to dec activity

Question 25

Bohr effect

Answer 25

Hemoglobin’s affinity to O2 based on pH and CO2; its affinity to O2 dec when low pH and high CO2

Question 26

Eqn for catalytic efficiency

Answer 26

Kcat/Km

Question 27

How to shift right on oxyhemoglobin dissociation curve?

Answer 27

Exercise is the right thing to do: increase [H+], increase PaCO2, increase temp, increase 2,3 bisphosphoglycerate