Control of Enzyme Activity (BIO, BC)

Question 1

General Catalysis

What is the function of an enzyme?

What will happen to the enzyme when we have an increased about of [S]

Answer 1

• Enzymes lower the activation energy of a reaction, or the ΔG of the transition state (NOT OF THE RXN!)
• E + S –>
• At really high [S] the enzymes will be saturated
  
  • Even if you increase concentration of [S] from this point, there will still be a Vmax

Question 2

Vmax

Answer 2

• is defined for a specific enzyme concentration (adding more enzyme will increase the Vmax)

Question 3

Michaelis-Menten equation

Answer 3

• calculates the rate of reaction using Vmax, the substrate concentration [S], and the Michaelis constant Km.
• Km = the [S] required to reach 1/2Vmax.
• As substrate concentration increases, the reaction rate also increases until a maximum value is reached
• At ½ Vmax, [S] = Km

Question 4

K_m

Answer 4

• Km does not fluctuate with changes in [enzyme] and is indicative of enzyme-substrate affinity
• Enzymes with high enzyme-substrate affinity will reach 1/2Vmax at a lower substrate concentration (Lower Km)
• Lower enzyme-substrate affinities will result in needing a higher substrate concentration to reach 1/2Vmax (Higher Km)

Question 5

K_cat

Answer 5

• = Enzyme’s “Turnover Number”
• How many substrates can this enzyme turn into product in one second at its maximum speed

Question 6

Catalytic Efficiency

Answer 6

Question 7

Kd & affinity

Answer 7

• Enzyme has lower Kd value—–> higher affinity for its substrate

Question 8

Cooperativity

Answer 8

• Some proteins can bind more than 1 substrate
• Cooperativity = substrate binding changes substrate affinity

Question 9

Positive, negative & non-cooperative

Answer 9

• Positive Cooperative Binding = Substrate binding increases affinity for subsequent substrate
• Negative Cooperative Binding = Substrate binding decreases affinity for subsequent substrate
• Non-Cooperative Binding = Substrate binding does not affect affinity for subsequent substrate

Question 10

Hemoglobin affinity for O2

T-state

R-state

Answer 10

“TOW RIGH”

• T state = Low affinity
• R state = High affinity

Question 11

Feedback Regulation

Answer 11

• When product of reaction binds allosteric site of the enzyme, affecting the catalytic activity
• Can be positive = increases enzyme-substrate affinity
• Can be inhibitory = reducing activity at the active site or inactivating it completely

Question 12

Competitive Inhibition

Answer 12

• E (inhibitor binds to E here to make EI) + S 🡨🡪 ES 🡨🡪 E + P
• Blocks the enzyme and makes it unable to react with substrate to form product
• Inhibitor competes with substrate for space on the enzyme
• Binds: Active Site
• Impact on Km: Increases
• Impact on Vmax: No Change

Question 13

Uncompetitive Inhibition

Answer 13

• E + S 🡨🡪 ES (inhibitor binds to the ES here to make ESI) 🡨🡪 E + P
• Molecule that binds only to the enzyme-substrate complex, rendering it catalytically inactive
• Binds: Allosteric Site
• Impact on Km: Decreases
• Impact on Vmax: Decreases

Question 14

Non-competitive

Answer 14

• Prevents the enzyme from turning substrate into product
• Binds to an allosteric site on the enzyme, causing a conformational change that decreases catalytic activity at the active site regardless of whether a substrate is already bound
• Binds: Allosteric Site
• Impact on Km: No Change
• Impact on Vmax: Decreases

***Bind the enzyme and the enzyme–substrate complex with the same affinity**

Question 15

Mixed inhibitor

Answer 15

• Molecule that binds to an allosteric site on the enzyme, causing a conformational change that decreases catalytic activity at the active site
• Generally, have preference towards binding either the enzyme-substrate complex, or binding the enzyme alone
• Binds: Allosteric Site
• Impact on Km: Increase (if prefer enzyme w/o substrate) or Decrease (if prefer enzyme with substrate bound)
• Impact on Vmax: Decreases

Question 16

Image of the line-burk plot of the inhibitors

Answer 16

Question 17

Allosteric Enzymes

Answer 17

Allosteric site present, molecule binds it, can either upregulate or downregulate the enzyme function

Question 18

Covalently-modified enzymes

methylation, acetylation, glycosylation & suicide inhibition

Answer 18

• Not all enzymes are proteins (i.e. Inorganic metals, small organic molecules like Flavin).

Small Posttranslational Modifications:

• Translation in synthesis of AA polymer
• “Post-translation” is after initial synthesis
• “Small” is adding or removing small functional groups

Methylation

• Modification of a protein that involves addition of methyl group (CH3)

Acetylation

• Modification of a protein that involves addition of an acetyl group

Glycosylation

• Addition of a sugar to a protein
  
  • I.e. Acetylation of lysine residue on a protein
  • Electron withdrawing impact of the acetyl group will prevent nitrogen from carrying positive charge and modify the behavior of the amino acid

Suicide Inhibition

• Suicide inhibitors covalently bind the enzyme and prevent it from catalyzing reactions
• Rarely unbind – why it’s called suicide (enzyme won’t work anymore)

Question 19

Zymogens

Answer 19

• Inactive form of an enzyme that requires covalent modification to become active
  
  • I.e. Digestive enzymes of the pancreas
    
    • Pancreas releases trypsinogen (a zymogen)
    • Once in the intestine, it is covalently modified by an enzyme called enterokinase to the active form Trypsin
    • This makes sure trypsin does not break down proteins that we need in the pancreas

Question 20

Hill coefficient

Answer 20

• Hill coefficient > 1, which means it exhibits cooperativity.
• Hill coefficient =1, no change for affinity of substrate
• Hill coefficient <1, negative cooperativity