Week 2 Enzymes

Question 1

Recap: Enzymes are biological catalysts

Answer 1

What does a catalyst do?
Increases the rate of a reaction, without being consumed by the reaction.

How does it do it?
By lowering the activation energy.

Question 2

Burning” glucose for fuel: Cellular respiration

Question 3

Enzymes in the body

Answer 3

Lots of digestive enzymes break down food.

Releases nutrients that make up our bodies and energy we need to exercise.
Enzymes catalyse just about every reaction that happens in cells.

Question 4

Enzymes in industry and agriculture

Answer 4

Rennet is a mix of enzymes used in cheesemaking: Pepsin breaks down proteins, Chymosin curdles milk, Lipases break down fats

Invertase breaks down hard sugars into soft sugars – used to make soft-centred chocolates

Phytases added to animal feed make minerals more bioavailable.

Question 5

What are enzymes and why are they necessary?

Answer 5

Enzymes are usually proteins and their activity depends on the integrity of their three-dimensional structure.

A denatured or dissociated enzyme usually won’t work.

Some enzymes require nothing other than their amino acids to function, but many enzymes require cofactors and/or coenzymes.

Uncatalysed reactions in biology tend to be slow—most biological molecules are quite stable in the neutral-pH, mild-temperature, aqueous environment inside cells.

Many biological reactions are chemically unfavourable so need enzymes.

Enzymes are usually much better (specific and efficient) than catalysts in chemistry.

If human enzymes don’t work properly it can lead to disease.

Question 6

Ribozymes: RNA enzymes

Answer 6

RNA molecules can have complex tertiary structures and so can be catalytically active.

Example: Hammerhead ribozymes promote site- specific cleavage of RNA molecules.

The RNA world hypothesis: very early in the evolutionary history of life RNA acted as both a storage molecule for genetic information and a catalyst for reactions.

Question 7

Not all enzymes exclusively use amino acid functional groups

Answer 7

Amino acids, with their various R-groups, can take part in lots of chemical reactions.

But some enzymes need extra molecules to function, these are often coenzymes and cofactors.

Question 8

Cofactors: metal ions

Answer 8

Cofactors are extra molecules or ions that are needed for the activity of some enzymes.
Very commonly these are metal ions like Fe2+ or Zn2+.

They are often found in enzymes that catalyse redox reactions (recall that Oxidation is loss of electrons, Reduction is gain of electrons). They also affect bonding, and can have a structural role.

More complex organic cofactors are called coenzymes.

The Zn2+ cofactor in carboxypeptidase A stabilises negatively charged groups in the amino acids surrounding it.

Question 9

Many metal ions are used as enzyme cofactors

Question 10

Coenzymes

Answer 10

Coenzymes are complex organic molecules needed for enzyme function.
They are often derived from vitamins.

They often act as transient carriers for functional groups.

Nicotinamide Adenine Dinucleotide (NAD) is very commonly used in redox reactions: it carries a hydride ion (:H-) and exists in reduced (NADH) and oxidised (NAD+) forms.

Question 11

Many types of coenzymes exist

Question 12

Some enzymes need both a coenzyme and a metal cofactor

Answer 12

Example: Alcohol dehydrogenase

Question 13

Prosthetic groups

Answer 13

Prosthetic groups (like haem) bind tightly or covalently to a protein, and act as structural elements. What we commonly call coenzymes bind more loosely. There are some inconsistencies in usage of the terms.

Cofactors-coenzymes. - prosthetic groups, cosubstrates
- metal ions.

Question 14

Holoenzymes and apoenzymes

Answer 14

Apoenzyme. Coezyme =. Substrate
(Protein portion) + Cofactor Holoenzyme
Inactive (Nonprotein portion). Whole enzyme
Activator Active

Question 15

Classifying enzymes

Answer 15

Many enzymes’ names end in –ase. E.g. urease breaks down urea.
Others have names derived from their roles. E.g. pepsin comes from the Greek word for digestion.

Some have several names or ambiguous names, so an international agreement has been made to sort enzymes into classes and sub-classes.

Question 16

Enzymes have active sites

Answer 16

An enzyme-catalysed reaction takes place in a pocket of the enzyme called the active site.

A substrate molecule binds in the active site and is acted upon by the enzyme.

The surface of the active site is lined with amino acid residues that allow the substrate to bind and catalyse its chemical transformation.

Often, the active site encloses a substrate, sequestering it completely from solution.

Question 17

Enzymatic reactions: Overview

Answer 17

A simple enzymatic reaction might be written:
E + S ⇌ ES ⇌ EP ⇌ E + P
Where E = enzyme, S = substrate, P = product.
Enzymes affect the rate of a reaction but do not affect its equilibrium. i.e.

They speed up the conversion of S→P they don’t make more P.

What’s important in enzymatic reactions is energy, in particular Gibbs Free Energy (G).
Recall: G = H – TS

(Gibbs free energy combines enthalpy, temperature and entropy).

Normally we compare the difference in free energy between systems, we write this as ΔG.

Question 18

Energy in reactions (uncatalysed)

Answer 18

As reaction progresses substrates (S) are converted into products (P). This leads to a change in Gibbs free energy of the system (ΔG).

In reactions old bonds are broken (uses energy) and new bonds are formed (gives out energy), S and P are different energy levels.

The energy level of “resting” S or P is called the ground state.
There is an activation energy barrier (ΔG‡) to overcome before the reaction can occur.

The transition state (‡) is the activated form of the molecules at the top of the “energy hill”, at this precise point decay back to S or progress to P is equally likely.

The higher the activation energy, the slower the reaction.

Question 19

Enzymes lower activation energy

Answer 19

In enzymatic reactions intermediates are introduced: Enzyme-substrate complex (ES) and enzyme-product complex (EP).

The activation energies required to produce these intermediates is lower and the transition state is at a lower energy level, so the rate of the reaction increases.
ΔG‡cat is lower than ΔG‡uncat.
Enzymes are not used up in the reaction, which is why we call them catalysts.

Question 20

Enzyme activity is affected by pH

Answer 20

If an enzyme is outside its optimum pH range, the amino acid side-chains will be incorrectly ionised leading to loss of function.
Pepsin is an endopeptidase, it hydrolyses peptide bonds to break down proteins into smaller peptides. It is found in the stomach.
Glucose 6-phosphatase hydrolyses glucose 6-phosphate to produce glucose and free phosphate. It is found in the liver.

Question 21

Cofactor

Answer 21

An inorganic ion or a coenzyme required for enzyme activity.

Question 22

Coenzyme

Answer 22

An organic cofactor required for the action of certain enzymes; often has a vitamin component.

Question 23

Prosthetic group

Answer 23

A metal ion or organic compound (other than an amino acid) covalently bound to a protein and essential to its activity.

Question 24

Holoenzyme

Answer 24

A catalytically active enzyme, including all necessary subunits, prosthetic groups, and cofactors.

Question 25

Apoenzyme

Answer 25

The protein portion of an enzyme, exclusive of any organic or inorganic cofactors or prosthetic groups that might be required for catalytic activity.

Question 26

Classifying enzymes

Answer 26

Many enzymes’ names end in –ase. E.g. urease breaks down urea.

Others have names derived from their roles. E.g. pepsin comes from the Greek word for digestion.

Some have several names or ambiguous names, so an international agreement has been made to sort enzymes into classes and sub-classes.

Question 27

Enzymes have active sites

Answer 27

An enzyme-catalysed reaction takes place in a pocket of the enzyme called the active site.

A substrate molecule binds in the active site and is acted upon by the enzyme.

The surface of the active site is lined with amino acid residues that allow the substrate to bind and catalyse its chemical transformation.

Often, the active site encloses a substrate, sequestering it completely from solution.

Question 28

Chymotrypsin is an example of an enzyme

Answer 28

Chymotrypsin is a protease that catalyses the hydrolysis of peptide bonds. It is specific for peptide bonds next to aromatic amino acids (Trp, Phe, Tyr).

Primary structure: 3 peptide chains linked by disulphide bridges. Important residues are highlighted.

Polypeptide backbone showing α-helices and β- sheets.n

Space-filling model. Binding pocket is yellow. Residues in active site are red.

Close-up of active site with substrate bound.

Question 29

Enzymatic reactions: Overview

Answer 29

A simple enzymatic reaction might be written:

E + S ⇌ ES ⇌ EP ⇌ E + P
Where E = enzyme, S = substrate, P = product.

Enzymes affect the rate of a reaction but do not affect its equilibrium
. i.e. They speed up the conversion of S→P they don’t make more P.

What’s important in enzymatic reactions is energy, in particular Gibbs Free Energy (G).

Recall: G = H – TS

(Gibbs free energy combines enthalpy, temperature and entropy).

Normally we compare the difference in free energy between systems, we write this as ΔG.

Question 30

Energy in reactions (uncatalysed)

Answer 30

As reaction progresses substrates (S) are converted into products (P). This leads to a change in Gibbs free energy of the system (ΔG).

In reactions old bonds are broken (uses energy) and new bonds are formed (gives out energy), S and P are different energy levels.

The energy level of “resting” S or P is called the ground state. There is an activation energy barrier (ΔG‡) to overcome before the reaction can occur.

The transition state (‡) is the activated form of the molecules at the top of the “energy hill”, at this precise point decay back to S or progress to P is equally likely.

The higher the activation energy, the slower the reaction.

Question 31

Enzymes lower activation energy

Answer 31

In enzymatic reactions intermediates are introduced: Enzyme-substrate complex (ES) and enzyme-product complex (EP).

The activation energies required to produce these intermediates is lower and the transition state is at a lower energy level, so the rate of the reaction increases.

ΔG‡cat is lower than ΔG‡uncat.

Enzymes are not used up in the reaction, which is why we call them catalysts.

Question 32

Enzyme activity is affected by pH

Answer 32

If an enzyme is outside its optimum pH range, the amino acid side-chains will be incorrectly ionised leading to loss of function.

Pepsin is an endopeptidase, it hydrolyses peptide bonds to break down proteins into smaller peptides. It is found in the stomach.

Glucose 6-phosphatase hydrolyses glucose 6-phosphate to produce glucose and free phosphate. It is found in the liver.

Question 33

Cofactor

Answer 33

An inorganic ion or a coenzyme required for enzyme activity.

Question 34

Coenzyme

Answer 34

An organic cofactor required for the action of certain enzymes; often has a vitamin component.

Question 35

Prosthetic group

Answer 35

A metal ion or organic compound (other than an amino acid) covalently bound to a protein and essential to its activity.

Question 36

Holoenzyme

Answer 36

A catalytically active enzyme, including all necessary subunits, prosthetic groups, and cofactors.

Question 37

Apoenzyme

Answer 37

The protein portion of an enzyme, exclusive of any organic or inorganic cofactors or prosthetic groups that might be required for catalytic activity.