Module 6 - Enzymes: function, reaction mechanisms, and kinetics

Question 1

How do enzyme active sites work?

Answer 1

provide chemical environments that facilitate catalytic reactions by excluding excess solvent (such as water)

Question 2

What are the three parameters of enzyme function?

Answer 2

enzymes bind substrates with high affinity and specificity, binding to active sites induce structural changes, and enzyme activity is highly regulated in cells (to balance catabolic and anabolic pathways)

Question 3

How is the catalytic efficiency of glycogen phosphorylase increased?

Answer 3

noncovalent binding of allosteric regulators like AMP and covalent attachment of a phosphoryl group on Ser14

Question 4

How do enzymes effect the conversion of hydrogen peroxide to water and oxygen?

Answer 4

half life of H2O2 would be 3 years, but by adding a free iron the half life is only 11.6 minutes

Question 5

How does catalase increase reaction rate?

Answer 5

assist in proton movement and oxidation of Fe3+ to Fe4+

catalase reaction is 10,000,000 more efficient than Fe alone

Question 6

How do catalysts and enzymes effect reaction energy?

Answer 6

lower the transition state

Question 7

What are cofactors vs. coenzymes?

Answer 7

cofactors are typically metal ions (Cu2+, Zn, etc.), while coenzymes are larger vitamin derivatives (NAD, FAD, CoA, etc.)

Question 8

What kind of cofactor does nitrite reductase have?

Answer 8

A Cu2+ ion cofactor between two His residues that hold it in an optimal position for catalyzing the reaction

Question 9

What is lipoamide?

Answer 9

a coenzyme that is covalently attached to a lysine residue in the decarboxylase enzyme

Question 10

What are the three ways that enzymes increase reaction rate?

Answer 10

stabilize the transition state, provide an alternate path for product formation via stable intermediates, and optimal substrate orientation and increased local concentration

Question 11

Uncatalyzed reactions require collisions that are…

Answer 11

at the right orientation and sufficient energy

Question 12

What are the two main functions of enzyme active sites?

Answer 12

• provide optimal orientation
• exclude excess solvent (water) that can interfere with the reaction

Question 13

What is demonstrated by the aldolase reaction?

Answer 13

the importance of favorable spatial arrangements, and the formation of a covalent substrate-enzyme intermediate

Question 14

In hexokinase, what happens at the active site?

Answer 14

when glucose binds, water is excluded to prevent nonproductive phosphoryl transfer from ATP to water

Question 15

How do hydrophobic substrate channels work?

Answer 15

prevent water from entering the buried active site

Question 16

What is the difference between specific and non specific acid base catalysis?

Answer 16

specific - involves water
non specific - proton transfer involving a functional group

Question 17

RNA cleavage with pancreatic ribonuclease involves…

Answer 17

two histidine residues and a general acid base catalysis using the addition of water

Question 18

What is the unique aspect of covalent catalysis?

Answer 18

a transient covalent bond forms between the substrate and the enzyme that can also be used for create an unstable intermediate

the other reactant will react more strongly with the unstable enzyme intermediate than it would with the original reactants

Question 19

What is an example of covalent catalysis?

Answer 19

1,3 BPG formation using the coenzyme NAD+

Question 20

What is an example of metal ion catalysis?

Answer 20

the carbonic anhydrase reaction uses a Zn 2+ ion as a catalytic group to yield the bicarbonate product

Question 21

What is the difference between redox reactions at C-O bonds vs. C-C bonds?

Answer 21

C-O: NAD+/NADH
C-C: FAD/FADH2
both transfer 2 electrons

Question 22

What are the three types of metabolite transformations?

Answer 22

isomerization:
reactions do not change the molecular formula of the product compared to that of the substrate

condensation:
reactions combine two substrates to form a larger molecule

hydrolysis or hydration: hydration or dehydration using water

Question 23

What are two examples of reversible modifications?

Answer 23

insulin signaling stimulates phosphoinositide-3-kinase activity which is turned off when phosphatase and tensin homolog removes the phosphate

DNA methyltransferases use the metabolite adenosyl-I-methionine as a methyl donor to add a methyl to deoxycytidine. The demethylation of deoxycytidine by a DNA glycosylase is associated with gene activation.

Question 24

Describe chymotrypsin reactions

Answer 24

Chymotrypsin is a serine protease that cleaves proteins using acid-base and covalent catalysis. Serine, histidine, and aspartate form a catalytic triad.

Consists of three peptide chains, with serine on the C chain and histidine and aspartate on the B chain.

Question 25

Describe enolase reactions

Answer 25

Enolase is a metalloenzyme with each active site containing two divalent metal ions that are required for the reaction. Lys acts as a general base and Glu acts as a general acid, overall resulting in the formation of phosphoenolpyruvate (PEP)

Question 26

Describe HMG-CoA reductase

Answer 26

tetrameric enzyme in the cholesterol biosynthetic pathway

atorvastatin is a cholesterol lowering drug

two hydride transfers involving two molecules of NADPH coenzyme with the active site stabilizing the transition state

Question 27

What are statin drugs?

Answer 27

treatments for artherosclerosis by lowering LDL levels

all are similar to HMG-CoA with different functional groups

Question 28

What does the rate constant, k, of a reaction indicate?

Answer 28

how quickly a substrate molecule is converted to product under defined conditions

Question 29

What is the equation for velocity of the reaction?

Answer 29

v=k[S] or v=k[S][Y]
v is amount of product formed per unit time, k is rate constant, and [S] and [Y] are concentration of substrates

Question 30

In Michaelis-Menten Kinetics, how is substrate concentration held “constant”?

Answer 30

the concentration of substrate is much greater than the concentration of enzyme, so it stays relatively the same

Question 31

What are the three assumption of Michaelis-Menten Kinetics?

Answer 31

• no product has been generated
• product released is assumed to be a rapid step
• concentration of ES remains approximately constant because [S]»>[E]

Question 32

What is Km? What does it mean about an enzyme?

Answer 32

the Michaelis constant, or the concentration of substrate at which velocity is 1/2 Vmax

low value means high catalytic activity

Question 33

What is the Michaelis-Menten equation? What does it represent?

Answer 33

Vo = Vmax [S] / Km + [S]

represent the hyperbolic curve of initial velocity and substrate concentration

Question 34

What is the Lineweaver-Burk plot?

Answer 34

graph that can be used to obtain the Michaelis-Menten parameters

slope = Km/Vmax
x intercept = -1/Km (same for all substrate concentrations)
y intercept = 1/Vmax

Question 35

What is kcat?

Answer 35

the turnover number, or the catalytic rate

Question 36

What value is best used for describing how well the enzyme works?

Answer 36

kcat/Km, the specificity constant

Question 37

How does pH and temperature affect enzyme activity?

Answer 37

Changes in pH can affect chemistry of enzymes because seven amino acids have ionizable side chains that could gain or lose a proton

Changes in temperature can affect catalytic properties, but also structure of the protein