Enzymes & Kinetics

Question 1

What are some general properties of enzymes?

Answer 1

Enzymes are proteins with catalytic activity which increase the rate of a chemical reaction without being consumed; exhibit stereospecificity.

Question 2

What is the energy of activation in a reaction?

Answer 2

The amount of energy it takes for a reaction to occur.

Question 3

What are the six enzyme classifications?

Answer 3

(1) Oxidoreductases
(2) Transferases
(3) Hydrolases
(4) Lyases
(5) Isomerases
(6) Ligases

Question 4

What are some characteristics of isoenzymes?

Answer 4

They catalyze the same reaction as enzymes, but they can exist in a different form. They can exist due to different amino acid composition, different physical properties, and different enzymatic properties.

Question 5

Complete the following equation:

_____ (active form) = _____ (inactive form) + _____

Answer 5

Holoenzyme Activity = Apoenzyme Activity + Cofactor

Question 6

Define cofactor(s).

Answer 6

non-protein molecule conjugated to an enzyme.

Question 7

What are examples of cofactors?

Answer 7

Essential ions, coenzymes, inorganic and organic, Ca, Cl-, Mg+, NADH, NADPH.

Digestive Enzymes.

Question 8

Define Prosthetic Group.

Answer 8

A tightly bound cofactor. E.g. heme bound to peroxidase.

Question 9

What are factors influencing enzyme activity?

Answer 9

Temperature, pH, Activators (cofactors), inhibitors, substrate concentration, and enzyme concentration.

Question 10

How does the temperature of the reaction effect the activity?

Answer 10

As temperature increases, activity will increase until the optimum temperature is reached, then denaturation of the protein will occur.

Question 11

How is activity of an enzyme reaction affected by activators and inhibitors?

Answer 11

Activators (cofactors) enhance the reaction (reaches max velocity the fastest).

Inhibitors delay the reaction (reaches the max velocity the slowest).

Question 12

How does the concentration of the substrate affect the enzyme reaction?

Answer 12

Reaction velocity is dependent on substrate concentration.

Question 13

During zero order kinetics, at saturating {S}, velocity is ___ of substrate concentration and ___ proportional to {E} enzyme concentration.

Answer 13

independent; directly

Question 14

Define competitive inhibitor.

Answer 14

Binds at the active site and competes with the substrate for binding sites.

Question 15

How is Km and Vmax altered during competitive inhibition?

Answer 15

Raises Km; Vmax is unaffected.

Question 16

What kind of inhibitor represents this graph?

Answer 16

Competitive Inhibitor.

Question 17

Define noncompetitive inhibitor.

Answer 17

Binds at an allosteric or regulatory site, which may be at or far removed from the active site.

Question 18

How is Km and Vmax altered during noncompetitive inhibition?

Answer 18

Km is unaffected; Vmax decreases.

Question 19

What kind of inhibitor represents this graph?

Answer 19

Noncompetitive Inhibitor

Question 20

Define uncompetitive inhibitor.

Answer 20

Binds to the enzyme-substrate complex and not to free enzyme; prevents product formation.

Question 21

How is Km and Vmax altered during uncompetitive inhibition?

Answer 21

Lowers Km; Lowers Vmax; Vmax/Km unchanged.

Question 22

What kind of inhibitor is represented by this graph?

Answer 22

Uncompetitive inhibitor.

Question 23

Define irreversible inhibition.

Answer 23

An inhibitor that binds specifically to the active site; forms stable covalent bond with enzyme inactivating it.

Question 24

Enzyme measurement is dependent on what?

Answer 24

Temperature and pH changes, activator concentration, inhibitor concentration, and substrate concentration.

Question 25

How are enzymes measured?

Answer 25

By activity, not concentration. It measures the rate or velocity.

Question 26

What is the international unit for measuring enzymes?

Answer 26

U: the amount of enzyme that catalyzes 1 umol of substrate per minute at standard conditions.

Question 27

NADH is measured at what wavelength?

Answer 27

340nm.

Question 28

What are the three phases of first order kinetics, and what do they do?

Answer 28

(1) Lag phase: E + S
(2) Linear phase, zero order: E + S = P; {S} is high and enzyme is saturated with substrate. Product formation is constant.
(3) Depletion phase: A plateau is reached; Vmax has been reached. The reaction rate reduces due to: ↓ [S], equilibrium of reaction, product inhibition, and pH changes.

Question 29

In zero order kinetics, when should enzyme activity be measured?

Answer 29

At saturating [S] (phase 2 - linear phase).

Question 30

In zero order kinetics, velocity is ___ of substrate concentration and ___ proportional to [E].

Answer 30

independent; directly

Question 31

When is there a deviation from linearity occur in enzyme reactions?

Answer 31

Due to high concentrations of enzymes, depletion of available substrate, and multiple readings of absorbance are missed because of the fast reaction rate.

Question 32

How can you solve deviation from linearity for an enzyme reaction?

Answer 32

By diluting the sample.

Question 33

Enzyme activity is calculated based on what?

Answer 33

Molar absorptivity of NADH, change in absorbance over time, volume, Beer’s law.

Question 34

Enzymatic reactions can be used to quantitate what analytes?

Answer 34

Glucose, cholesterol, and uric acid.

Question 35

Define absolute specificity.

Answer 35

One substrate, one reaction.

Question 36

Define group specificity.

Answer 36

Reactions with a certain class of molecules. 
E.g. ACP can hydrolyze beta glycerol phosphate.

Question 37

Define stereoisomeric specificity.

Answer 37

Specific for one optical isomer.

Question 38

How does sex alter specimens?

Answer 38

Differences related to muscle mass, exercise or hormone concentration.

Question 39

How does age alter specimens?

Answer 39

Variations occur during first year of life, puberty, and late to middle age.

Question 40

How does exercise CK-MM?

Answer 40

Increases.

Question 41

What analytes are affected by hemolysis?

Answer 41

LD, ALP, AMS, ALT, AST, GGT, ACP.