5-enzymes A Flashcards
whats a ligand
a bound molecule
what is the range of ligand types
from small molecules to other proteins
is ligand binding covalent or non covalent
non covalent
where does ligand binding occur
at the binding site which is complementary to ligand
what causes the ligand binding site to be complementary to the ligand (4, 4 (he has two slightly diff things on two slides)
hydrophobic interactions, van der waals, ionic interactions, h bonds
CHARGE H-BOND POLARITY SHAPE
is ligan binding cooperative or non cooperative
it can be both
is the curve like in non-cooperative ligand binding
it has a hyperbolic curve
is the curve like in cooperative ligand binding and why
sigmoidal binding curve (allostery)
what kind of curve for allostery
it is sigmoidal
what is Kd
the concentration of ligand to get 50% bound
what does a small Kd mean with affinity
greater the binding affinity of the ligand for its target
what does a large Kd mean with affinity
the more weakly the target molecule and ligand are attracted to and bind to one another
In presence of 5uM ni, a nickel binding protein is found to be 25% saturated with nickel (theta = 0.25). What is the value of Kd?
15uM
what do enzymes do to kinetic rate
increase
are enzymes specific
yes
what are most of enzymes made from
protein
what kind of conditions do most enzymes operate under
mild conditions (low temp and pressure)
what are some things that some enzymes require
cofactors
can enzymes be regulated
yes
what does the active site provide to enzymes
A favorable sequestered environment for biologically important reactions to occur
what are cofactors
additional chemical components that assist in enzyme function
what are 3 examples/types of cofactors
metal ions, coenzymes, prosthetic groups
what are prosthetic groups
metal ions or coenzymes that are tightly or covalently associated with a protein
what is a holoenzyme
enzyme+prosthetic group
what is an apoenzyme
enzyme without a prosthetic group
what do enzymes do to the activation energy of a reaction
reduce
what do enzymes do to the dela G of the whole reaction
nothing
what is deltadelta G
the reduction in deltaG by a catalyst
what is deltadetlaG often reffered to as
Binding Energy
what is binding energy
deltadeltaG of the catalyzed reaction
can there be more than one transition state and intermediate with reactions
yes
what determines the overall rate of the reaction
the rate limiting step
what are 2 things that the rate of reaction is related to
concentration of substrates and rate constate
what does an increase in T do to rate
increase
what does a modest change in activation energy do to rate
significant change (exponential)
what happens to k (rate constant) when you increase activation energy
it decreases
what are the 7 enzyme classes
- oxidoreductases
- transferases
- hydrolases
- lyases
- isomerases
- ligases
- translocases
what class of enzyme is composed of reduction reaction
oxidoreductases
what class of enzyme is composed of oxidation reactions
oxidoreductase
what do oxidoreductases do
enzymes that catalyse oxido-reductions
what kind of enzyme is dehydrogenases
oxidoreductases
what kind of enzyme is oxidases
oxidoreductases
what kind of enzyme is reductases
oxidoreductases
what class of enzyme is composed of a group transfer
transferases
what are transferases
enzymes that transfer a group to another compound
what kind of enzyme class is a kinase
transferase
what kind of enzymes catalyse hydrolysis reactions
hydrolases
what do hydrolases do
catalyse hydrolysis reactions
which enzyme class is often named with the suffix -ase
hydrolases
what kind of enzyme class is a protease
hydrolases
what kind of enzymes cleave C-C, C-N, C-O bonds but NOT via hydrolysis or oxidation
lyases
what do lyases do
cleave C-C, C-N, C-O bonds but NOT via hydrolysis or oxidation
what enzyme class is often named the synthases
lyases
what are a common name for lyases
synthases
what is special about lyase reactions
in one direction of the reaction, there is one compound fewer
what kind of enzyme family catalyses elimination and addition reactions
lyases
what do isomerases do
catalyze structural rearrangements within a single molecule
which kind of enzymes catalyze structural rearrangements within a single molecule
isomerases
what kind of enzymatic reaction would turn a ketone into an aldehyde
isomerases
what do ligases do
catalyse the joining of 2 molecules or 2 parts of a molecule at the expense of a H moneluce - ATP hydrolysis
what is the enzymatic reaction that catalyses the joining of 2 molecules or 2 parts of a molecule at the expense of a H moneluce - ATP hydrolysis
ligases
what is required in a ligase reaction
a high energy molecule like ATP or GTP
what do translocases do
catalyse the movement of ions or molecules across membranes, or their seperation within membranes
what kind of enzyme catalyse the movement of ions or molecules across membranes, or their seperation within membranes
translocases
what kind of enzyme family would the Na+K+ ATPase pump be
translocase
once you know these, practice with the glycolysis diagram in the notes
okie dokie
what are 4 ways that enzymes lower the activation energy
participate in reaction
desolvation
proximity/orientation
stabolie transition state
what are 3 ways that enzymes participate in reactions to increase reaction rate
(Directly or indirectly)
- general acid/base catalysis
- nucleophilic/covalent catalysis
- metal ion catalysis
how does proximity and orientation reduce activation energy
reduces entropy
what is general acid base catalysis
proton transfer/donation to/from specific side chains in the enzyme active site
what kind of catalysis is RNase
acid base
what are the 2 ways that spontaneous alkaline hydrolysis can occur
which is more common
either a 2’ or 3’ nucleoside monophosphate
3’ is more common
what does His 12 and His 119 do in RNas acid base catalysis
His 12 is base His119 is acid
where do covalent bonds occur in non-covalent catalysis
between/within substrate molecules, not the enzyme
where do covalent bonds occur in covalent catalysis
enzyme becomes covalently attached to the intermediate
what does covalent catalysis do to the reaction pathway
alters the pathway!!
will the intermediates be same in a catalyzed and an uncatalyzed reaction
no
what are nucleophiles
anything with an appropriate lone pair
what are 2 examples of electrophiles
H+ and metal ions
what are 3 ways that metal ions can participate in catalysis
- binding/orienting of substrate
- stabilizing of transition state
- redox centers
what kind of catalasis happens with carbonic anhydrase
metalloenzyme –> metal ion catalysis
what happens to reaction rate when the two reactive groups are spatially constrained (close together)
the reaction rate increases
what are 2 ways to reduce entropy in a reaction
increase proximity and orientation
what is desolvation
substrate binding removing the water (remove water surrounding enviro so they can interaction)
why do you wanna do desolvation
when water forms a shell around many small molecules, you want to strip away the water so the groups can interact
what is induced fit and why does it happen
when enzmyes change shape upon substrate binding to bring catalytic groups into orientation
can induced fit be used to exclude water
yes it can
what kind of enzyme is hexokinase (which class and why)
transferase because it transfers a phosphate onto substrate using ATP
what is the hexokinase reaction a “classic example” of
induced fit
is induced fit with hexokinase covalent or non covalent catalysis
non-covalent
what does hexokinase do to the poshpate
enhances its electrophilicty
what do the histidines do in RNase
act as either a general base or a general acid
what enzyme class is RNase
hydrolase
does RNase do covalent cataylsis
no
what kind of enzyme class is carbonic anhydrase and why
lyase
reaction is H2CO3 - H2O + CO2
what enzyme class is PEP carboxykinase
transferase
what assisnts PEP carboxykinase
metal ion cofactors
what is the nucleophile in the hexokinase reaction and where do they attack
O (from C6)from the OH attacks the PO4-
what does hexokinase provide to the reaction / act as
a general base
what in hexokinase acts as a general base
Asp
what does Mg2+ do in the hexokinase mechanism
interacts with the negative charge
what enhances the difference in activation energies (between catalyzed and uncatalyzed)
the strong interactions with transition state
does strong interactions of enzymes with transition state enhances the difference in activation energies (between catalyzed and uncatalyzed)
yes
does strong interactions of enzymes with substrate enhances the difference in activation energies (between catalyzed and uncatalyzed)
no, only with transition state
what is the enzyme class of serine proteases
hydrolases
what kind of catalysis is serine protease (covalent or non)
covalent
what kind of catalysis is hexokinase (covalent or non)
non covalent
what kind of catalysis is RNase (covalent or non)
non covalent
what does serine proteases do
breaks a polypeptide chain, generates new N and C termina
what do serine proteases do to transition state
stabilize
what is a catalytic triad and which enzyme has this
3 residues that are critical in the reaction mechanisms, in serine proteases
which residues are in the catalytic triad of serine proteases
His 57
Ser 195
Asp 102
what does His 57 do
general acid/base
what does Ser 195 do
nucleophile
what does Asp 102 do
modulates His 57 (makes sure its in the right orientation and protonation state)
what are zygomens
inactive precursors
what activates zygomens
cleavage of the peptide chain
what are serine proteases synthesized as
zygomens
how are serine proteases activated
by cleavage of peptide chain
how many domains in serine proteases and what are thet
2 beta barrel domains
where is the active site in serine protease with respect to the domains
at the junction of the 2 beta barrel domains
where are serine proteases located in the cell
extracellular
do serine proteases have disulfides and why
yes because they are extracellular
what is the “specificity pocket” in serine protease
a part of the active site which interacts favourably with some aa and not others - determines substrate specificity (which ones will be cleaved) so like chymotrypsin vs. trypsin vs. elastase
which amino acid in chymotrypsin acts as the nucleophile
serine 195 (the OH group)
what kind of enzyme is chymotripsin
a serine protease (hydrolase)
which amino acid in chymotrypsin acts as the the general acid/base
His 57
which amino acid in chymotrypsin helps maintain orientation and protonation state of His 57
Asp 102
what protonation state do you want His 57 to be in in chymotrypsin what happens (how do you know if it is in the right state)
protonated
they will form an H bond with eachother
what does the oxyanion hole do (main role)
transition-state stabilization
what is a oxyanion hole
pocket in the active site of an enzyme that stabilizes transition state negative charge on a deprotonated oxygen
what kind of catalysis is chymotrypsin (covalent or non)
covalent
what does chymotrypsin like to cleave the most
cleave peptides after large hydrophobic residues
what does trypsin like to cleave the most
cleave peptides after positively charged residues
what does elastase like to cleave the most
cleave peptides after small non polar residues
is the specificity pocket part of the catalytic region of the active site
no
is the specificity pocket part of the active site
yes
which serine protease likes to cleave after large hydrophobic residues
chymotripsin
which serine protease likes to cleave after positively charged residues
trypsin
which serine protease likes to cleave after small non polar residues
elastate
what forms the oxyanion hole
backbone NH groups of residue 193 195
is the oxyanion hole partially negative or negative
positive
who can the oxyanion hole form hydrogen bond interactions with
negatively-charged oxygen that develops during the reaction
what happens in the 1st step of the chymotrypsin mechanism (2 things)
His 57 (general base) becomes protonated, ser 195 does nucleophilic attack on substrate polypeptide
what happens to the C=O on the substrate polypeptide after step 1 in the chymotrypsin mechanism
it becomes C-O- (negative)
in step one, is His 57 a general base or acid and why
general base because it becomes protonated
in step two, is His 57 a general base or acid and why
general acid because it loses its H+
what happens in the 2nd step of the chymotrypsin mechanism (3 things)
His57 acts as a general acid, N from peptide bond in substrate becomes protonated (thats the first product)
also ser 195 becomes an acyl intermediated
what happens in the 3rd step of the chymotrypsin mechanism (1 thing)
amine (first product) is released and replaced with water
what happens in the 4th step of the chymotrypsin mechanism (2 things)
His 57 is a general base and takes the H from H2O
water acts as a nucleophile and attacks the carbonyl
in step four, is His 57 a general base or acid and why
general base because it steals the H from water
what happens in the 5th step of the chymotrypsin mechanism (2 things)
His 57 acts as a general acid (releases its H+)
generate the 2nd product (the new C terminus) and your enzyme returns to its origional state
which step of the chymotripson mechanisms creats the new N-terminus
2
which step of the chymotripson mechanisms creats the new C-terminus
5
how/why does trypsin cleave after positively charged residues
it has an aspartate in the specificity pocket
what is the scissile bond
the bond that is gonna be cleaved
What role does a general base play in the mechanism of hexokinase?
It accepts a proton during nucleophilic attack of a phosphate.
