Enzymes and Kinetics Flashcards

1
Q

What is a general way to describe how enzymes work?

A

The substrate binds to the active site, forms the enzyme-substrate complex, then the product leaves

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2
Q

What is the active site?

A

A pocket in the surface of the enzyme formed from precise 3D arrangements of the amino acids and/or prosthetic groups

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3
Q

What does the thermodynamics tell us about a reaction?

A

If the reaction is energetically favourable, which side is favoured by the equilibrium, what is the final distribution of products and reactants

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4
Q

What does the kinetics tell us about a reaction?

A

How fast the reaction will get to that final distribution

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5
Q

If deltaG < 0:

a) endergonic or exergonic?
b) products or reactants favoured?

A

a) exergonic

b) products

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6
Q

If deltaG > 0:

a) endergonic or exergonic?
b) products or reactants favoured?

A

a) endergonic

b) reactants

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7
Q

Do enzymes change the thermodynamics or the kinetics?

A

Only the kinetics

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8
Q

What is the transition state?

A

The high energy state at the peak of the activation barrier. It has an equal chance of becoming the product or substrate

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9
Q

How do enzymes speed up the reaction rate?

A

They lower the activation energy and interact with the reactants so that they reach the transition state easier

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10
Q

Do enzymes change the final distribution of products and reactants?

A

No, that is governed by deltaG, which enzymes do not change

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11
Q

What are 4 ways enzymes lower the activation energy?

A
  1. Desolvation
  2. Participating in the reaction
  3. Proximity and orientation effects
  4. Selective stabilization of the transition state
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12
Q

What is desolvation? Why does this help speed up the reaction?

A

Removing the reactants from water. It helps cause an induced fit between binding partners and forms the functional active site. Water can also interfere with reactions

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13
Q

What are proximity and orientation effects? Why does this help speed up the reaction?

A

The enzyme holds the two substrates together in the right orientation. The reaction goes faster than if they were floating around and hoping to bump into each other in the right orientation

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14
Q

How does an enzyme participate in the reaction?

A

Can provide functional groups, act as a nucleophile, acid-base assistance, redox reactions, then get regenerated later

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15
Q

What is selective stabilization of the transition state? Why does this help speed up the reaction?

A

The active site forms additional favourable interactions with the transition state. This makes the transition state favourable to form and lowers its energy

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16
Q

What are cofactors?

A

Non amino acid groups that provide additional chemical functionality. Includes metal ions and coenzymes

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17
Q

What is the mechanism of enzymes?

A

Break up a large reaction into several smaller ones that still have a lower activation energy overall

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18
Q

What is the function of lysozymes?

A

Cleaves peptidoglycan so bacteria can’t withstand osmotic pressure

19
Q

What are the two important amino acids for the catalytic function of lysozyme?

A

Glu35 and Asp52

20
Q

What is different about Glu35 in lysozyme?

A

It is surrounded by hydrophobic amino acids. It is unfavourable for it to be in the A- form, so it stays in HA and has a much higher pKa

21
Q

What is the optimal pH for lysozyme?

A

5

22
Q

What is the mechanism by which lysozyme breaks down peptidoglycan?

A
  1. Asp52 acts as a nucleophile and attacks the glycosidic bond
  2. Forms a covalent bond with the peptidoglycan
  3. Glu35 protonates it and half of the product is released
  4. SN2 attack from water
  5. Glu35 deprotonates the water
  6. Breaks covalent bond with Asp52
  7. Second half of the product is released and regenerates the enzyme
23
Q

Why is enzyme kinetics useful?

A

Can tell us the reaction mechanism, how it changes with different enzymes and substrates, the functioning of inhibitors

24
Q

What shape is the Michaelis-Menten graph?

A

Hyperbolic

25
Q

What shape is a Lineweaver-Burk graph?

A

Linear

26
Q

What is on the x and y axis in the Michaelis-Menten graph?

A

x: substrate concentration
y: velocity

27
Q

What is the slope in a Michaelis-Menten graph?

A

At the beginning it is V0

28
Q

What is on the x and y axis in a Lineweaver-Burk graph?

A

x: 1/substrate concentration
y: 1/velocity

29
Q

What is the slope in a Lineweaver-Burk graph?

A

Km/Vmax

30
Q

What are 7 factors that can influence enzyme activity?

A

Substrate concentration, enzyme concentration, temperature, pH, inhibitors, allostery, regulation

31
Q

How can enzyme concentration affect enzyme activity?

A

Increases Vmax because there’s more active sites

32
Q

How do high temperatures affect enzyme activity?

A

Activity drops at high temperatures because the protein gets denatured

33
Q

How do low temperatures affect enzyme activity?

A

They lose their flexibility, which is important for function

34
Q

How does pH affect enzyme activity?

A

Outside of the narrow optimal pH range, amino acid side chains get protonated or deprotonated when they shouldn’t be

35
Q

What is an inhibitor?

A

A molecule that interferes with catalysis

36
Q

What interactions does a reversible inhibitor use to bind to its target?

A

Non-covalent interactions like hydrogen bonds

37
Q

What interactions does a irreversible inhibitor use to bind to its target?

A

Covalent bonds, permanent inactivation

38
Q

What are the 3 types of reversible inhibitors?

A

Competitive, uncompetitive, and mixed

39
Q

How can we determine the type of inhibitor a molecule is?

A

Experimentally. We measure the rate of reaction at different inhibitor concentrations and determine what effect it has on the kinetics

40
Q

What is a competitive inhibitor? Where does it bind?

A

An inhibitor that competes with the substrate, looks like it but doesn’t react. It binds in the active site

41
Q

How do competitive inhibitors affect the y-intercept, Vmax, slope, and Km on the Lineweaver-Burk graph?

A

The y-intercept and Vmax are unaffected. The slope and Km increase because more substrate is required to fill half the active sites

42
Q

How do uncompetitive inhibitors affect the y-intercept, Vmax, slope, and Km on the Lineweaver-Burk graph?

A

The y-intercept increases, Vmax decreases. Km decreases proportionally to Vmax. The slope is unaffected

43
Q

How do mixed inhibitors affect the y-intercept, Vmax, slope, and Km on the Lineweaver-Burk graph?

A

The y-intercept and slope increase. Vmax decreases. Km can either increase or decrease

44
Q

How can allostery affect enzyme activity?

A

By binding to the enzyme and changing its shape to be active (allosteric activator) or inactive (allosteric inhibitor)