Lecture 12 - Enzyme Kinetics

Question 1

How many different enzymes are there?

Answer 1

about 80,000

Question 2

What are the 6 different classes of enzymes?

Answer 2

oxidoreductases, transferases, hydrolases, lyases, isomerases, ligases

Question 3

1) Define oxidation-reduction rxns 2) Give an example and label the donor, acceptor, reduced substrate and oxidized substrate

Answer 3

1) transfer of electrosn (hydride ions or H+ atoms) from one substrate (donor) to a second substrate (acceptor) 2) Lactate –> pyruvate; lactate = donor and reduced substrate, NAD+ = acceptor and oxidized substrate

Question 4

Define oxidoreductases and give two examples

Answer 4

1) transfer of electrons (hydride ions or H atoms) 2) dehydrogenases, oxidases, reductases, oxygenases, hydroxylases

Question 5

Define transferases and give two examples

Answer 5

1) group transfer rxns 2) acyl, methyl, glucosyl and phosphoryl transferases; kinases, mutases

Question 6

Define hydrolases and give two examples

Answer 6

1) hydrolysis rxn - transfer of fun gps to water 2) esterases, glycosidases, peptidases, phosphatases, phospholipases, ribonucleases

Question 7

Define lyase and give two examples

Answer 7

1) addition of gps to dbl bonds or formation of dbl bonds by removal of groups 2) decarboxylases, aldolases, synthases, lyases

Question 8

Define isomerases and give two examples

Answer 8

transfer of gps within a molecules –> isomers 2) epimerases, isomerases

Question 9

Define ligases and give two examples

Answer 9

1) formation of C-X bonds where X=C, S, O, N; coupled to ATP cleavage 2) synthetases, carboxylases

Question 10

1) What is a phosphorylation rxn kinase an example of? 2) how many substrates and products do these rxns have?

Answer 10

transferases –> transfer of a chemical gp from one molecule to another 2) two substrates and two products

Question 11

Give another example of a transferase that we talked about in class and explain what happens

Answer 11

Transglutaminase –> form extensively cross-linked generally insoluble protein polymers (meat glue)

Question 12

What is a key point to remember about hydrolases when classifying?

give an example

Answer 12

they are NOT classified as transferases even though they can be thought of as transferring a fn gp to water; usually an IRREVERSIBLE RXN

Dephosphorylating Serine –> P gp is transferred to water –> HOPO (-2)

Question 13

What does Alpha-galactosidase do?

in what product can you find this enzyme?

Answer 13

breaks down complex (branching) sugars such as polysaccharides in food su ch as legumes and cruciferous vegs (broccoli, cauliflower) ; complex sugars –> simple sugars –> food somewhat more digestible

Beano

Question 14

What is an effect of Isomerases on the C atom of a molecule?

give an example of a rxn

Answer 14

May change the sterochemistry at a carbon atom within the molecule; moving a group or a dbl bond within the same molecule

D-Lactic acid –> L-lactic acid

Question 15

What is the common action of lyases and describe the two types

Answer 15

usually breaking apart of a C-C bond

synthase –> no ATP used

synthetase –> ATP used

Question 16

true or false:

Lyases are never reversible

Answer 16

False:

Some rxns are reversible

Question 17

What do Ligases require for the rxn to occur (joining C-C bonds)?

Answer 17

ATP (synthetase) = ATP –> ADP + Pi

Question 18

True or False:

1) Almost all enzymes are proteins.
2) most rxns in a cell don’t require the action of an enzyme
3) Enzymes act as catalysts.
4) An enzyme speeds the rate of rxn as well as changes the equilibria of a rxn

Answer 18

1) True. RNA is an enzyme that isn’t a protein
2) False. most rxns require an enzyme since most will not occur under the physical conditions in a cell (and if they do, the rxn will occur slowly)
3) True. They increase the rate of chemical rxns to convert substrate –> product
4) False. they speed up the rate of rxn but DO NOT alter the equilibria of a rxn

Question 19

How does an ezyme speed up the rate of rxn?

Answer 19

Energy is used to break and reform bonds (S–>P) so enzymes decrease the activation E (delta G)

Question 20

What is the activation E?

Answer 20

the amount of E needed to go from a ground state to a transition state

E+S <–> ES <–> EP <–> E+P

ES <–>EP is the transition state

Question 21

1) How do Enzymes lower delta G?
2) what type of interactinos are involved between the E and its S?
3) what do these interactions provide?
4) What promotes the formation of the transition state or E-S complex?

Answer 21

1) they provide an alternative, lower-energy rxn path
2) many weak noncovalent bonds (h-bonding)
3) Binding E –> major source of free energy used by E to lower activation energies of rxns
4) interaction of the E and the S at the active site

***provides a stable state for ES complex –> release of E –> lower activation E

Question 22

Describe the two theories of substrate binding to an enzyme

Answer 22

1) Lock and Key theory: the shape of the substrate fits the enzyme

E+S can’t get to transition state

2) Induced fit theory : the enzyme is flexible (they are dynamic, can move and bend)

enzyme more resembles transition state which allows the substrate to morph with enzyme into TS

Question 23

What is enzyme specificity?

Answer 23

ability of an enzyme to discriminate btw two completing substrates. Same binding E used for catalysis also makes the E specific

Question 24

Why are Enzymes so big when active sites are a relatively small part of its total volume?

Answer 24

scaffolding, regulatory sites, interaction sites for other proteins and channels; they are large because multiple weak interactions are requird to drive catalysis (stabilize structure; active site) –> helps form ES complex

Question 25

what do coenzymes and cofactors do?

what are they derived from?

example

Answer 25

help enzymes with rxns; coenzymes are regenerated at the end; cosubstrates can be used up

vitamins

biotin –> carboxylation

Question 26

What is this diagram an example of?

Answer 26

A reversible rxn and how substrate and product can reach equilibrium (rate of S = rate of P)

Question 27

What is this diagram indicative of?

Answer 27

1) As you saturate the enzyme, concentration of P will increase as well as the rate of rxn
2) Initial Velocity and amount of P (where you reach equilibria and Vmax) increases with increasing S concentrations
3) Eventually will reach equilibrium and max velocity –> after awhile, increasing substrates won’t have an effect since there are only so many E (constant #), enzymes are going as fast as they can –> even if dbl enzymes, will still eventually reach max velocity

Question 28

what is zero order kinetics

Answer 28

when substrate and enzyme are equally balanced

Question 29

Describe this diagram

Answer 29

1) Rxn will never go faster than the rate
2) As you increaes the E concentration, you increase initial velocity ( 4 X E = 4 X Vo)
3) As you increase E, increase Vmax, as you decrease E, you decrease Vmax

Question 30

Which step is slowest in going from E–>P?

Answer 30

K2 (ES–> E+P)

Vo = most of E is not bound to S

Vmax = most of E is in ES c omplex

Question 31

1) What is Km?
2) what does Km depend on

Answer 31

1) Km = Vmax/2
2) depends upon rate constants and overall raxn –> useful indicator of how well a substrate interacts with an enzyme (how tightly something binds)

how well you stabilize rxn

NOT indicative of the affinity of E for S

Question 32

1) What can changes in Km values be used for?
2) How can mutations impact Km values?
3) give a clinical correlation

Answer 32

1) Changes are useful in comparing enzyme functino
2) mutations can affect structure of enzyme –> structure of substrate binding site –> Km –> rxn rate at typical S concentrations
3) ALDH removes acetaldehyde generated by alcohol dehydrogenase by converting acetaldehyde to acetate (acetaldehyde is highly toxic metabolite). Altered Km affects the rate of removal of acetaldehyde

Hangover = didn’t convert a lot of acetaldehyde to acetate

Question 33

Name this reaction

Answer 33

oxidation -reduction

Question 34

Name this reaction

Answer 34

Phosphorylation reaction kinases (involving transferases)

Question 35

Name this reaction

Answer 35

hydrolysis involving hydrolase

Question 36

Name this reaction

Answer 36

isomerization onvolving isomerases

Question 37

Name this reaction

Answer 37

lyase reaction

Question 38

Name this reaction

Answer 38

ligase reaction