Introduction to Enzyme Kinetics

Question 1

How do enzymes speed up the rate of reactions?

Answer 1

By lowering the free energy activation for a reaction. They do not change the standard free energy of the product

Question 2

First order reaction

Answer 2

• Reaction rate is directly proportional to substrate concentration
• Enzyme kinetics follow this pattern prior to reaching Vmax

Question 3

Zero order reaction

Answer 3

• The maximum velocity of a reaction is reached and active sites are almost continuously filled
• Increase in substrate concentration after this point will not increase the rate

Question 4

Michaelis constant (Km)

Answer 4

Substrate concentration required to reach one half of Vmax

At this [S], half of the enzymes are bound to S

Question 5

How are Km and Enzyme/Substrate affinity related?

Answer 5

They are inversily related

Higher Km = lower affinity

Lower Km = higher affinity

Question 6

The enzyme velocity is at one-half the max rate when what percent of the enzyme is bound to substrate?

Answer 6

50% of the enzyme is bound to substrate

Question 7

What happens to the Vmax when more enzyme is added?

Answer 7

The Vmax increases

Question 8

What happens to the Vmax when the enzyme concentration is reduced by 50%?

Answer 8

The Vmax is reduced by 50%

Question 9

What does Kcat represent?

Answer 9

The turnover number which describes how many substrate molecules are transformed into products per unit time by a single enzyme

Question 10

Lineweaver-Burk plot

Answer 10

• X-axis = 1/[S]
• X-intercept = 1/Km
• Y-axis = 1/Vo
• Y-intercept = 1/Vmax
• Slope = Km/Vmax

Question 11

What are the 4 main types of enzyme inhibition?

Answer 11

• Competitive inhibition
• Non-competitive inhibition
• Irreversible inhibition
• Allosteric enzyme

Question 12

Competitive inhibition

Answer 12

• Competes with a substrate for binding at the enzyme’s substrate-recognition site
• Vmax = unchanged
  
  • Competitive inhibitors can be overcome by increasing the substrate concentration
• Km = increased
  
  • More substrate is needed to achhieve 1/2 Vmax

Question 13

Competitive inhibition on plots

Answer 13

Question 14

Noncompetitive inhibition

Answer 14

• Inhibitor and substrate bind at different sites
• Inhibitor can bind E or ES complex and prevents the reaction from occuring
• Vmax = Decreases Vmax
  
  • Cannot be overcome with additional substrate
• Km = unchanged
  
  • Noncompetitive inhibitors do not interfere with the binding of substrate thus do not change the affinity

Question 15

Noncompetitive inhibition plots

Answer 15

Question 16

Irreversible inhibition

Answer 16

• Structural analog of the substrate is converted to a more effective inhibitor with the help of the enzyme to be inhibited
• New product irreversibly binds to the enzyme and inhibits further reaction
• If [inhibitor] > [enzyme] –> No reactions as all enzymes are inhibited

Question 17

What are three main factors that influence enzymatic activity?

Answer 17

• Temperature: rate of reaction increases with temperature then decreases as enzymes denature at high temperatures
  
  • Most animal enzymes rapidly denature above 40 C
• pH: Different enzymes have different optimal pHs
  
  • Some enzymes are only function at high or low pHs
• Substrate concentration: increases in [S] increase velocity up to the point of Vmax at which point more S no longer increases the velocity

Question 18

Allosteric enzymes

Answer 18

• Regulated by molecules called effectors that noncovalently bind at a site other than the active site
• Composed of multiple subunits and regulatory (allosteric) site that bind effector
• Frequently catalyze commited step early in a pathway

Question 19

Effector

Answer 19

Molecule that regulates allosteric enzymes by noncovalently binding at a site other than the active site

Question 20

Positive effector

Answer 20

Either increases the Vmax or decreases the Km

Question 21

Negative effector

Answer 21

Either decreases Vmax or increases Km

Question 22

Homotropic effector

Answer 22

• When the substrate iteself serves as an effector
• Often (but not always) are positive effectors, where the presence of substrate molecule at one site enhances the catalytic properties of the other site
  
  • This is called cooperativity –> Exhibits a sigmoidal curve

Question 23

Heterotropic effector

Answer 23

The effector is different from the substrate

Question 24

What type of allosteric effect does carbon dioxide have on hemoglobin’s ability to carry oxygen?

Answer 24

CO2 acts as a negative, heterotropic, allosteric effector

Question 25

What step are regulatory enzymes usually involved in?

Answer 25

The rate-limiting step or the commited step