enzymes mw%% +

Question 1

Active site complementary to substrate = no reaction!

Answer 1

Question 2

Explain the allosteric control of enzymes?

Answer 2

• Allosteric effectors are cell metabolites, bind non-covalently to a site on the enzyme that is not the active site
• This changes the enzymes structure
• Some effectors are activators and some are inhibitors
• Allosteric effectors are examples of non-competitive inhibitors

Question 3

Explain Covalent Modification?

Answer 3

•Many ways enzymes (and other proteins) can be regulated through reversible covalent modification e.g phosphorylation.

Question 4

What is an Enzyme?

Answer 4

• Globular protein
• Biological catalyst that differs from a chemical catalyst

–Catalyses very high reaction rates

–Shows great reaction specificity

–Work in mild temperature/pH conditions

–Can be regulated

Question 5

Enzyme Definitions?

Answer 5

• Apoenzyme–The protein component of an enzyme that contains a cofactor
• Holoenzyme –“whole enzyme” – the apoenzyme plus the cofactor(s)

Question 6

why don’t Allosteric enzymes not follow M-M kinetics?

Answer 6

• Allosteric enzymes are made up of many subunits, which contain many active sites.
• One substrate binding to an enzyme subunit can cause changes in other active sites on other subunits
• This can lead to the concept of “cooperative binding” of substrate molecules

Question 7

what is a coenzyme?

Answer 7

• Complex organic molecule, usually produced from a vitamin e.g NAD

Question 8

What do Enzymes do?

Answer 8

They do NOT:

–Move reaction equilibria

–Make a non-spontaneous reaction spontaneous

They DO:

–Increase rates of spontaneous reactions

–Lower the activation energy of biochemical reactions

–Accelerate movement towards reaction equilibrium

Question 9

What will affect an enzyme?

Answer 9

Temperature

• ↑ temp., ↑ molecule collisions
• ↑ temp., ↑ internal energy of molecules
• ↑ temp. will eventually denature enzyme

pH

• pH changes the charge of amino acids
• If the active site amino acids charge changes the enzyme will cease to function correctly

Inhibitors

Question 10

Reaction Coordinate Diagram?

Answer 10

• Spontaneous reaction isn’t instantaneous because of the energy barrier
• Transition state shows the moment that chemical bonds are formed and broken
• Reaction could then go either way (revert back to S or change to P)

Question 11

what is a free energy?

Answer 11

• Energy can be converted from one form to another but is neither created or destroyed.
• Useful” energy generated from cellular reactions is termed Gibbs Free-Energy (G).
• Spontaneous reactions must have a –ve ΔG value

Question 12

how do irreversible inhibitors affect enzymes?

Answer 12

inhibitor binds to the enzyme in a covalent, and therefore irreversible way.

Question 13

what is a Prosthetic group?

Answer 13

• Cofactor covalently bound to the enzyme or very tightly associated with the enzyme

Question 14

Measuring enzyme activity

Answer 14

•In clinical samples, normal activity is often given an arbitrary value:

1 U/ml or 100% = normal

•Separate different forms of enzymes by electrophoresis and examine pattern

Question 15

Enzymes and two or more substrates?

Answer 15

• Catalysing a reaction with two or more substrates usually involves transfer of groups from one substrate to the other
• This can occur in several ways:

–Random order or Ordered with a ternary complex (complex formed between two substrate molecules and an enzyme)

–No ternary complex formation

Question 16

How can you measure enzyme kinetics?

Answer 16

• At low [S] you get an almost linear increase in V₀ as ↑[S]
• At higher [S] the V₀ changes very little in response to ↑[S]
• When [S] becomes so large that V₀ changes are vanishingly small you get maximum reaction velocity, V_max
• Vmax occurs because all of the enzyme active sites are saturated with substrate

Question 17

what are Isoenzymes?

Answer 17

• Different enzymes that catalyse the same reaction e.g hexokinase (muscle) glucokinase (liver)

Question 18

how do non-competitive inhibitors affect enzymes?

Answer 18

• bind to enzymes non-covalently
• The substrate is still able to bind the active site, so Km of the substrate-enzyme complex remains unchanged
• Increasing substrate concentration does not change the inhibition, Vmax will decrease
• non-competitive inhibitors= unchanged Km values but the Vmax decreases.

Question 19

how do Competitive inhibitors affect enzymes?

Answer 19

• bind to enzymes non-covalently ,resemble the substrate molecule,compete with the active site
• leads to a decrease in the affinity between the active site and the substrate, so the Km increases
• Increasing substrate concentration can overcome this inhibition, so the same Vmax can be achieved
• competitive inhibitors exhibit increased Km values, Vmax remains unchanged

Question 20

what are the functions of Glucokinase and hexokinase?

Answer 20

They both catalyse:

• Glucose + ATP ⇒ Glucose-6-phosphate + ADP.
• When [blood glucose] goes UP, the glucokinase activity increases but hexokinase activity does not respond as it is already working at its Vmax
• When [blood glucose] is LOW, gluconeogensis releases glucose from the liver,

Question 21

how does feedback inhibition occur?

Answer 21

• Some biochemical pathways regulate the enzymes involved through feedback inhibition.
• A build up of the end product of a pathway, or a key junction in a pathway, can ultimately slow the entire pathway

Question 22

What can Km and Vmax tell us?

Answer 22

• Km gives you a clue to the affinity of the enzyme with it’s substrate.
• larger Km values indicate a less stable ES complex. Reduced affinity
• smaller Km values indicate a more stable ES complex. Higher affinity
• Vmax tells you how fast a reaction is proceeding when the enzyme is saturated with substrate

Question 23

How do enzymes reduce activation energy?

Answer 23

Entropy reduction

–Molecules in free solution will only react by “bumping” into one another

–Enzymes “force” the substrate(s) to be correctly orientated by binding them in the formation they need to be in for the reaction to proceed

Desolvation

–Weak bonds between the substrate and enzyme replace H-bonds between substrate and aqueous solution

Induced fit

–Conformational changes occur in the protein structure when the substrate binds

Question 24

what is a cofactor?

Answer 24

• Non-protein component needed for activity

Question 25

what are the enzymes that catalyse the phosphorylation & dephosphorylation of enzymes?

Answer 25

* Protein **kinases** – **add** phosphoryl groups to proteins * Protein **phosphatases** – **remove** phosphoryl groups

Question 26

Explain Proteolytic Cleavage?

Answer 26

* Enzymes can exist as an **inactive precursor protein**, called a proprotein or proenzyme. * Proproteins can be **cleaved** to give **active enzyme by proteases**

Question 27

Active site complementary to transition state = CATALYSIS

Answer 27

Question 28

Enzyme Reaction Coordinate Diagram?

Answer 28

* An enzyme will **facilitate** the change of S to P through intermediates. * Enzymes form n**on-covalent bonds** with substrate molecules, called the “binding energy” allowing them to take the reaction through a different path of reaction intermediates

Question 29

how is enzyme activity controlled in cells?

Answer 29

•Two main ways **regulatory enzymes** modulate reactions: 1. **Allosteric** enzymes 2. **Covalently modified** enzymes •Both classes of regulatory enzymes tend to be **multi-subunit** proteins