Lecture 10: HOW DOES AN ENZYME CATALYSE A REACTION? 1

Question 1

What are the strategies for catalysis?

Answer 1

Acid-base catalysis, covalent catalysis, redox and radical catalysis, geometric effects, stabilisation of the transition state and cofactors with activated groups

Question 2

What is redox and radical catalysis driven by?

Answer 2

Metal ions

Question 3

What are the geometric effects?

Answer 3

Proximity and orientation

Question 4

what are some examples of activated groups of cofactors?

Answer 4

Electrons, hydride ions (H-), methyl groups (CH3), amino groups (NH3) with the ability to carry things

Question 5

What do many enzymes do?

Answer 5

Use more than one strategy

Question 6

What does covalent catalysis involve?

Answer 6

The formation of a reactive, short-lived intermediate, which is covalently attached to the enzyme

Question 7

What does the enzyme in covalent catalysis have?

Answer 7

A nucleophilic side chain

Question 8

How is the enzyme regenerated in covalent catalysis?

Answer 8

By a hydrolysis reaction

Question 9

What is covalent catalysis driven by?

Answer 9

Nucleophilic attack on an electrophile

Question 10

For two molecules to react they need to be …

Answer 10

Close together and in the correct orientation

Question 11

What does nucleophilic attack require?

Answer 11

Correct orientation and ionisation

Question 12

What does nucleophilic attack depend on?

Answer 12

The R groups and how good the leaving group is

Question 13

What does nucleophilic attack form?

Answer 13

A trigonal planar intermediate

Question 14

What should allow nucleophilic attack to occur faster?

Answer 14

Stabilising the transition state

Question 15

What does hexokinase use as a cofactor?

Answer 15

Mg2+

Question 16

What is established in hexokinase?

Answer 16

Orientation of phosphates of ATP by octahedral coordination of Mg2+ ion

Question 17

What is the Mg2+ cofactor known as?

Answer 17

An electron withdrawing Lewis acid which stabilises oxygen, making phosphorus a better electrophile (leaving group)

Question 18

What does acid-base catalysis involve?

Answer 18

Ionisable groups and proton transfer

Question 19

What amino acids have ionisable groups?

Answer 19

Glu, Asp (COOH)

Lys, Arg (NH3+)

Question 20

What do amino acid groups need to be in for the catalytic mechanism to proceed?

Answer 20

In the correct ionisation state

Question 21

What may ionisation be part of?

Answer 21

activation to the transition state

Question 22

What is enzyme activity dependent on?

Answer 22

pH

Question 23

Why is enzyme activity ph dependent?

Answer 23

Because transfer of protons is involved

Question 24

What does each enzyme have?

Answer 24

A characteristic optimal pH at which its rate is highest

Question 25

What is histidine particularly suitable to?

Answer 25

Acid-base catalysis reactions

Question 26

What is the pKa of the imidazole group in histidine?

Answer 26

~6.5 which is close to physiological pH

Question 27

What can histidine do?

Answer 27

Depending on the environment of the active site His can donate or accept a proton

Question 28

What does chymotrypsin do?

Answer 28

Cleave proteins

Question 29

What catalysis strategies does chymotrypsin use?

Answer 29

Acid-base, covalent, geometric effects and stabilisation of the transition state

Question 30

What are proteases?

Answer 30

Hydrolyases

Question 31

What are the substrates in a reaction using chymotrypsin?

Answer 31

A polypeptide and water

Question 32

What are the products in a reaction using chymotrypsin?

Answer 32

Two shorter peptides, or individual amino acids

Question 33

What does chymotrypsin act in?

Answer 33

Digestion

Question 34

What do other proteases do?

Answer 34

Make hormones, remove defunct proteins we no longer need, or activate enzymes (as in blood clotting)

Question 35

What do divergent evolution serine proteases have?

Answer 35

Same structure with unique specificities

Question 36

What is in the specificity pocket of chymotrypsin?

Answer 36

2 glycines (small)

Question 37

What is in the specificity pocket of trypsin?

Answer 37

Negative aspartic acid which attracts positively charged polypeptides

Question 38

What is in the specificity pocket of elastase?

Answer 38

Valine and threonine with large side chains

Question 39

What do convergent serine proteases have?

Answer 39

The same catalytic triad occurs in different order and different structures.

Question 40

What should proteases have if they shared a common ancestor?

Answer 40

Share order and structure

Question 41

What does the catalytic triad in chymotrypsin include?

Answer 41

Serine, histidine and aspartic acid

Question 42

What is the first part in the action of chymotrypsin?

Answer 42

his57, polarised by Asp102, withdraws a proton from Ser195 making there serine hydroxyl a good nucleophile to attack the sissile bond

Question 43

What is the sissile bond?

Answer 43

The bond which is broken

Question 44

What happens after his57, polarised by Asp102, withdraws a proton from Ser195 making there serine hydroxyl a good nucleophile to attack the sissile bond?

Answer 44

Nucleophilic attack forms a covalent, tetrahedral intermediate which is stabilised by the oxyanion hole, lowering the activation energy

Question 45

what happens after Nucleophilic attack forms a covalent, tetrahedral intermediate which is stabilised by the oxyanion whole, lowering the activation energy?

Answer 45

The tetrahedral intermediate decomposes, driven by general acid catalysis from his57

Question 46

What happens as a result of the first tetrahedral intermediate decomposing?

Answer 46

Half of the polypeptide chain remains covalently attached to the enzyme in an acyl-enzyme intermediate and the other half can leave the active site

Question 47

What happens after the first half leaves the active site?

Answer 47

Water replaces one of the products in the active site

Question 48

What happens after water replaces one of the products in the active site?

Answer 48

Polarised by His57 acting as a general base (OH-), water makes a nucleophilic attack forming a second tetrahedral intermediate, again stabilised by the oxyanion hole

Question 49

What happens after Polarised by His57 acting as a general base (OH-), water makes a nucleophilic attack forming a second tetrahedral intermediate, again stabilised by the oxyanion hole?

Answer 49

Whit His57 acting as a general acid, this intermediate decays to form a carboxylate

Question 50

What happens as a result of the second intermediate decaying to form a carboxylate?

Answer 50

The second product can leave the active site and the active site has been regenerated and is ready for another round of catalysis