Lecture 8

Question 1

Standard chemical reactions consist of?

Answer 1

• Reactant molecules on one side
• Product molecules on one side
• Transition state between the two

Question 2

standard chemical reaction?

Answer 2

A population of molecules with a distribution of energy.

Question 3

Spontaneous direction of standard chemical reaction?

Answer 3

Spontaneous direction corresponds to delta G. But the speed at which products go to reactants or reactants go to products is dependent on having reactant molecules that have a high enough free energy to get over the free energy barrier.

Question 4

Transition state?

Answer 4

The transition state is a barrier to chemical reactions, as it is very high and hard for reactants and products to make it across.

Question 5

What do catalysts do?

Answer 5

Catalysis lowers the activation energy for a reaction, lowering the free energy barrier.

• More molecules are able to reach the new delta G, allowing more molecules to react.
• Catalysts increase reaction rates by decreasing delta G

Question 6

what does a lower delta G do?

Answer 6

A lower delta G allows more molecules to react.

Question 7

Kinetic energy and product formation?

Answer 7

Only molecules with enoguh kinetic energy can form product.

Question 8

Temperature and reaction rates?

Answer 8

Chance of forming product is lower with low temperature, and higher temperatures allow more molecules to react.

Question 9

Pros and cons to higher temperatures?

Answer 9

Pros: speeds up the reaction and leaves more energy for the actual reaction.

Cons: There are trade-offs - including denatured proteins, cell death, melting. Therefore, varying temperatures in a cell is a very bad idea.

Question 10

What factors can increase the reaction rate?

Answer 10

Increasing number of molecules, increasing the temperature,
or adding a catalyst will increase reaction rates.

Question 11

What are enzymes?

Answer 11

Enzymes are protein catalysts (except for ribozymes).

Question 12

Facts about enzymes?

Answer 12

Are the largest group of proteins (six families).
* Do not change the reaction equilibrium (i.e. catalyze reaction in both directions)
* Are not chemically changed by the reaction
* Are normally present in low amounts relative
to the reactants and products

Question 13

How do enzymes work?

Answer 13

Have active sites that bind substrate/product. Put substrates in proximity, with correct orientation and provide functional groups for catalysis (decrease activation energy). Preferentially bind and stabilize the transition state (induced fit)

Question 14

How effective are enzymes?

Answer 14

Extremely effective! They can enhance rates up to 10^17 fold! Typically, they only increase reaction rates by 10^8 - 10^12 fold (up to 10^6 reactions per second, or turnover time is 1 microsecond)

Question 15

Induced fit model?

Answer 15

The enzyme stabilizes the transition state:

Substrate binds into the enzyme. The enzyme binding the substrate puts it in a bound and stabilized position where it cannot move. Sometimes it stabilizes a transition state in which the substrate would be more likely to form products (p1 and p2 might come together to fit into the binding portion of the enzyme).

Question 16

What are cofactors?

Answer 16

Cofactors are molecules that provide additional chemistries for carrying out catalysis.

Question 17

Are cofactors required for all enzymes?

Answer 17

No, but they are required for many enzumes to carry out enzymatic activity.

Question 18

Apo-enzyme vs. holo-enzymes?

Answer 18

Apo-enzyme: without cofactor
Holo-enzyme: with cofactor

Question 19

Branches of Cofacotrs?

Answer 19

Metal ion: typically transition metal
Coenzymes: organic molecules - two kinds

Cosubstrates: Loosely bound such as NADH and NAD+
Prosthetic group: Tightly bound such as heme.

Question 20

Enzyme catalytic mechainisms>?

Answer 20

1. Acid base
2. Covalent catalysis
3. Metal catalysis

Question 21

Acid-base catalysis?

Answer 21

transfer or
removal of H+

Question 22

Covalent catalysis?

Answer 22

transient
formation of a covalent bond
between enzyme and substrateMetal

Question 23

Metal catalysis?

Answer 23

direct or indirect
role in catalysis; often oxidation-
reduction Rx

Question 24

Enzymes have…?

Answer 24

Enzymes have characteristic pH and temperature optima.

Question 25

Nucleophiles?

Answer 25

Nucleophiles (electron-rich and attracted to nuclei). Nucleophiles attack electron-deficient centers to create covalent bonds. Ex. Serine, tyrosine, Cysteine

Question 26

Electrophiles?

Answer 26

Electrophiles (electron-poor and attracted to electrons) Ex. H, Na, Ca, Carbonyl

Question 27

Metal Ions as Catalysts?

Answer 27

Metal ions can mediate oxidation-reduction reactions, promote reactivity of the substrate or other functional groups.

Question 28

Alcohol Dehydrogenase?

Answer 28

Zn stabilizes O during transfer from H (from NADH)

Question 29

Enzymes function by stabilizing A. their substrates B. their products C. the transition state D. their substrates and the transition state E. all of the above

Answer 29

C. The transition state By stabilizing the transiotin state, enzymes can reduce the activation energy by allowing the very unstable transition state between reactants and products to be stabilized, so that the reactants can become the products.

Question 30

Example: Chymotrypsin?

Answer 30

There is a peptide bond that needs to be broken for us to digest food. This reaction is catalyzed by chymotrypsin. There are 3 parts of chymotrypsin: - Asp 102 (stabilizes His57) - His57 (acid-base catalysis) - Ser195 (serine protease - significant)

Question 31

Chymotrypsin uses..?

Answer 31

Chymotrypsin uses both covalent and acid-base catalysis to hydrolyze a peptide bond (enhancement of ~ 1010).

Question 32

Step 1 of chymotrypsin?

Answer 32

His accepts a proton from Ser (base catalysis) allowing the nucleophilic oxygen of serine to attack the carbonyl carbon of the substrate (covalent catalysis). Asp102 stabilizes His57

Question 33

Step 2 of chymotrypsin?

Answer 33

His donates a proton to the new NH2 of the C-terminal peptide fragment, breaking the peptide bond. This forms the tetrahedral intermediate transition state.

Question 34

Step 3 of chymotrypsin?

Answer 34

Departure of the Rc leaves the enzyme covalently bound to the RN region of the substrate. Forms Acyl-enzyme intermediate (s stable covalent intermediate)

Question 35

Step 4 of chymotrypsin?

Answer 35

Water donates a proton to His 57 and resulting OH- attacks the carbonyl group of substrate (resembles step 1).

Question 36

Step 5 of chymotrypsin?

Answer 36

His 57 donating a proton leads to collapse of the second tetrahedral intermediate (resembles step 2). this forms a second tetrehedral intermediate.

Question 37

Step 6 of chymotrypsin?

Answer 37

N-terminal portion of substrate diffuses away, regenerating chymotrypsin. Chymotrypsin returns to its original state.

Question 38

In the following step of chymotrypsin catalysis (H-O bond goes to the C) A. His 57 is undergoing acid catalysis B. Ser 195 is undergoing base catalysis C. Asp 102 is undergoing acid catalysis D. Ser 195 is undergoing covalent catalysis E. None of the above

Answer 38

D. Ser 195 is undergoing covalent catalysis.

Question 39

Chymotrypsin catalysis: other factors?

Answer 39

- Even with mutation of Asp 102, His 57, and Ser 195, proximity and orientation still enhance the reaction rate by 10^5. - Gly 193 and a backbone amide stabilizes the transition state (Mutants with Ser 195 replaced are at least 106 fold less active, Mutants with Asp 102 or His 57 replaced are 1,000 fold less active)

Question 40

Divergent evolution of chymotrypsin?

Answer 40

It has evolved several times over - inside chymotrypsin, trypsin, and elastase. - Different substrates are recognized in related enzymes due to different binding pocket complementarity. - The substrate binding pocket complements the substrate.

Question 41

Catalytic Triads..?

Answer 41

Catalytic triads are used throughout biology.

Question 42

Bacterial substilisin?

Answer 42

Subtilisin has the same catalytic triad, but in a completely unrelated protein. This is an example of convergent evolution

Question 43

Isozymes?

Answer 43

Two different enzymes that catalyze the same reaction.