Lecture 12 - Enzyme Kinetics Flashcards
How many different enzymes are there?
about 80,000
What are the 6 different classes of enzymes?
oxidoreductases, transferases, hydrolases, lyases, isomerases, ligases
1) Define oxidation-reduction rxns 2) Give an example and label the donor, acceptor, reduced substrate and oxidized substrate
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
Define oxidoreductases and give two examples
1) transfer of electrons (hydride ions or H atoms) 2) dehydrogenases, oxidases, reductases, oxygenases, hydroxylases
Define transferases and give two examples
1) group transfer rxns 2) acyl, methyl, glucosyl and phosphoryl transferases; kinases, mutases
Define hydrolases and give two examples
1) hydrolysis rxn - transfer of fun gps to water 2) esterases, glycosidases, peptidases, phosphatases, phospholipases, ribonucleases
Define lyase and give two examples
1) addition of gps to dbl bonds or formation of dbl bonds by removal of groups 2) decarboxylases, aldolases, synthases, lyases
Define isomerases and give two examples
transfer of gps within a molecules –> isomers 2) epimerases, isomerases
Define ligases and give two examples
1) formation of C-X bonds where X=C, S, O, N; coupled to ATP cleavage 2) synthetases, carboxylases
1) What is a phosphorylation rxn kinase an example of? 2) how many substrates and products do these rxns have?
transferases –> transfer of a chemical gp from one molecule to another 2) two substrates and two products
Give another example of a transferase that we talked about in class and explain what happens
Transglutaminase –> form extensively cross-linked generally insoluble protein polymers (meat glue)
What is a key point to remember about hydrolases when classifying?
give an example
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)
What does Alpha-galactosidase do?
in what product can you find this enzyme?
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
What is an effect of Isomerases on the C atom of a molecule?
give an example of a rxn
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
What is the common action of lyases and describe the two types
usually breaking apart of a C-C bond
synthase –> no ATP used
synthetase –> ATP used
true or false:
Lyases are never reversible
False:
Some rxns are reversible
What do Ligases require for the rxn to occur (joining C-C bonds)?
ATP (synthetase) = ATP –> ADP + Pi
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
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
How does an ezyme speed up the rate of rxn?
Energy is used to break and reform bonds (S–>P) so enzymes decrease the activation E (delta G)
What is the activation E?
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
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?
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
Describe the two theories of substrate binding to an enzyme
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
What is enzyme specificity?
ability of an enzyme to discriminate btw two completing substrates. Same binding E used for catalysis also makes the E specific
Why are Enzymes so big when active sites are a relatively small part of its total volume?
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
what do coenzymes and cofactors do?
what are they derived from?
example
help enzymes with rxns; coenzymes are regenerated at the end; cosubstrates can be used up
vitamins
biotin –> carboxylation
What is this diagram an example of?
A reversible rxn and how substrate and product can reach equilibrium (rate of S = rate of P)
What is this diagram indicative of?
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
what is zero order kinetics
when substrate and enzyme are equally balanced
Describe this diagram
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
Which step is slowest in going from E–>P?
K2 (ES–> E+P)
Vo = most of E is not bound to S
Vmax = most of E is in ES c omplex
1) What is Km?
2) what does Km depend on
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
1) What can changes in Km values be used for?
2) How can mutations impact Km values?
3) give a clinical correlation
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
Name this reaction
oxidation -reduction
Name this reaction
Phosphorylation reaction kinases (involving transferases)
Name this reaction
hydrolysis involving hydrolase
Name this reaction
isomerization onvolving isomerases
Name this reaction
lyase reaction
Name this reaction
ligase reaction
