Lecture 13. Enzymes and Catalysis

Question 1

What does Eyring’s equation show ?

Answer 1

The rate of reaction can be related to Gibbs free energy of the transition state for that reaction

Question 2

What is the activation state ?

Answer 2

The energy required to get to the transition state from starting materials

Question 3

What happens as the activation energy increases ?

Answer 3

The rate constant (k) becomes smaller

The reaction becomes slower

Question 4

What can catalysts do ?

Answer 4

1. Reduce the activation energy for a reaction

2. Provide an alternative mechanism with a lower overall activation energy

Question 5

Why are enzymes crucial for life ?

Answer 5

Activation energies for uncatalysed metabolic processes are so big that very high temperatures would be requires to achieve the same rates

Question 6

What does a large difference in the activation energy result in ?

Answer 6

A larger difference in k catalysed and k uncatalysed

Question 7

What can enzymes provide ?

Answer 7

A large reduction in the activation energy

Question 8

What are the four categories by which enzymes catalyse reactions ?

Answer 8

1. Proximity and orientation effects
2. Acid base catalysis
3. Covalent catalysis
4. Transition state stabilisation

Question 9

What type of cost does binding a substrate to an enzyme require ?

Answer 9

Entropic cost

Question 10

What is required for a substrate to bind tightly to an enzyme ?

Answer 10

Favourable enthalpy

Question 11

What does binding of substrate molecules to enzymes allow ?

Answer 11

The enzyme to exert control over the conformation and shape of the substrate molecule

Question 12

What does free rotation around single bonds in acyclic systems result in ?

Answer 12

A huge number of thermally accessible conformations

Question 13

Which type of reaction (intramolecular or intermolecular) are generally faster ?

Answer 13

Intramolecular

Question 14

Why are intramolecular reactions generally faster ?

Answer 14

The reactants are already in the molecule

Question 15

What can contribute to influencing the rate of acceleration ?

Answer 15

1. Different functionality from different amino groups
2. Shape and form of protein
3. Interactions with the amide backbone

Question 16

In bimolecular reaction what are proximity and orientation effects thought to account for ?

Answer 16

Accelerations up to 10^8 fold over the background uncatalysed reaction

Question 17

How can protonation affect functional groups in substrate molecules ?

Answer 17

Turn them into better leaving groups

Question 18

How does protonation affect carbonyl groups ?

Answer 18

Makes them more electrophilic

Question 19

What can deprotonation do to functional groups ?

Answer 19

Make them into better nucleophiles

Question 20

What does acid/base catalysis do by appropriate amino acid side chains ?

Answer 20

Accelerating enzymatic processes

Question 21

What amino acid cannot be chiral ?

Answer 21

Glycine

Question 22

What amino acids have carboxylic side chains ?

Answer 22

Aspartic acid and glutamic acid

Question 23

What happens if aspartic acid and glutamic acid are protonated in the active sites ?

Answer 23

They can act as proton donors

Question 24

What happens if aspartic acid and glutamic acid are deprotonated ?

Answer 24

They can stabilise adjacent positive charges and perhaps act as bases (proton acceptors)

Question 25

What happens when the pH of an environment corresponds to the molecules pKa ?

Answer 25

The compound will be 50% ionised

Question 26

What can nucleophilic sides chains react directly with and form ?

Answer 26

Substrates forming covalent bonds between the enzyme and the substrate

Question 27

What happens to the covalent bonds formed between the enzyme and substrate ?

Answer 27

Cleaved in later steps, regenerating the unmodified enzyme

Question 28

What can lysine react with ?

Answer 28

Carbonyl groups in substrates to form imines/iminium ions

Question 29

What does thioester formation occur with ?

Answer 29

Cysteine side chains

Question 30

What can favorable reactions between the enzyme and substrate do ?

Answer 30

Stabilise the transition state and lower the activation energy

Question 31

What are some methods of understanding how an enzyme works ?

Answer 31

1. Kinetic studies
2. Detection of intermediates
3. Chemical modification

Question 32

What is x-ray crystallography used for?

Answer 32

A very powerful technique that allows for the 3D structure of the enzyme to be solved

Question 33

How would you obtain the structure of the enzyme-substrate complex ?

Answer 33

Grow crystals and soak in substrates

Question 34

What is the problem of growing and soaking the crystals in substrate ?

Answer 34

Enzyme turnover is generally much faster than the timescale of crystallisation and data aquisition

Question 35

How would you solve the problem of a quick enzyme turnover in x-ray crystallography ?

Answer 35

1. Use an unreactive substate mimic
2. If the enzyme requires two substrates, try two crystals one with each substrate
3. Complex of the enzyme with the product

Question 36

What can the use of x-ray crystallography in conjunction with computational modelling do ?

Answer 36

Reverse some of the tricks that are used to obtain crystal structures

Question 37

What is site directed mutagenesis ?

Answer 37

Amino acids are exchanged for others

Question 38

Which amino acids should you mutate ?

Answer 38

Target key structural/catalytical residues

Question 39

In site directed mutagenesis, how would you make as little disruption as possible to the enzyme structure ?

Answer 39

1. Remove the reactive group

2. Replace with something of similar size (ideally smaller) and polarity/charge distribution