Enzymes (EK B1 Ch1) COPY Flashcards
oxidoreductases
reaction catalyzed= transfer of hydrogen and oxygen atoms or electrons from one substrate to another ex. dehyrogenases, oxidases, oxidation reduction reactions
Oxidoreductases (including dehydrogenases) catalyze redox reactions
transferases
catalyze reactions of the transfer of a specific group (a phosphate or methyl etc.) from one substrate to another ex. transaminase, kinase so groups are transferred from one location to another
hydrolases
regulate hydrolysis of a substrate / hydrolysis reactions ex. estrases, digestive enzymes
isomerases
change of the molecular form of the substrate/ transfer of groups within a molecule, with the effect of producing isomers ex. phospho hexo, isomarse, fumarase
lysases
nonhydrolytic removal of a group or addition of a group to a substrate
- so functional groups are added to double bonds or conversely, double bonds are formed via the removal of functional groups
ex. decarboxylases, aldolases
Lyases break covalent bonds using mechanisms besides hydrolysis
ligases (sythetases)
- joining of two molecules by the formation of new bonds
- catalyze condensation reactions coupled with hydrolysis of high energy moelcules
ex. citric acid synthetase
hexokinase
- lowers activation energy for the phosphorlyation of glucose
- enzyme that phosphorylates glucose as soon as it enters the cell
from metabolism ch1 (Ch 4 in entire packet)
Step 1: glucose is phosphorylated to glucose-6-phosphate by hexokinase
- ATP is the source of the phosphate group
- Glucose is trapped in the cell. Could go on through glycolysis or be stored as glycogen
- Hexokinase is in most cells, including muscle and brain
- Hexokinase has low Km (high affinity for glucose) but low Vmax
- Hexokinase will work even when glucose levels are low
saturation kinetics
as relative concentration of substrate inc, the rate of reaction also increases, but to a lesser and lesser degree until a maximum rate, Vmax has been achieved. = this occurs because as more substrate is added, individual substrates must begin to wait in line for an unoccupied enzyme
saturation kinetics workers ex
analogous to assemebly line workers -when there are more workers, more enzymes, the rate of production VMAX increases! -but there comes a point when there is just too much starting material (substrate) and the workers cannot go any faster -at this point the workers or enzymes are saturated and have reached Vmax
vmax
proportional to enzyme concentration
kcat
turnover number
- number of substrate molecules one active site can convert to product in a given unit of time when an enzyme solution is saturated with substrate
- provides a rough estimate of the catalytic efficiency of an enzyme
kcat
= vmax/Et (vmax/ enzyme concentration)
Km
= 1/2 vmax, it is the substrate concentration at which the reaction rate equals to 1/2 vmax
Km explanation
indicates how highly concentrated the substrate must be to speed up the reaction
- if a higher concentration of substrate is needed, the enzyme must have a LOWER AFFINITY for the substrate
- Km is inversely proportional to the enzyme substrate affinity****
- does not vary when the enzyme concentration is changed, unlike vmax** in other words, it is characteristic of the intrinsic fit between the enzyme and substrate, rather than reflecting amount fo substrate present
glucokinase
this and hexokinase add a phosphate to glucose, to form glucose-6-phosphate, which is trapped inside the cell
- has a significantly higher Km, meaning that it has lower affinity for glucose compared to hexokinase
- acts in liver, so high levels of blood glucose would be required to begin phosphorylating glucose in the heptocyte cytosol
- this lower affinity allows glucose to be phosphorylated in other cells, which use hexokinase!!! Only when glucose concentrations become high will the liver begin storing it as glycogen and fatty acids
temp affects rates of enzymatic reactions
as temp inc, the reaction rate initially goes up
- since enzymes are generally proteins, at some point the enzyme denatures and the rate of reaction drops off precipitously
- for enzymes in human body, optimal temp is around 37 C
pepsin
enzyme in stomach prefers pH of 2
trypsin
enzyme active in small intestine, works best at a pH btw 6 and 7
cofactor
for optimal activity some enzymes need cofactors to function -either minerals or coenzymes -nonprotein component
Enzyme regulation: 4 primary means
- proteolytic cleavage (irreversible covalent modification), ex zymogen
- reversible covalent modification, ex. amp, protein kinase when some enzymes activated or deactivated by phosphorylation or the addition of some other modifier such as AMP, removal of modifier is always accomplished by hydrolysis
- control proteins- protein subunits that associate with certain enzymes to activate or inhibit their activity, ex calmodulin and g proteins
- allosteric interactions
zymogen
many enzymes released into their environment in the INACTIVE FORM called a zymogen or proenzyme (greek pro= before) when specific peptide bonds or zymogens are cleaved, the zymogen become irreversibly activated. activation of zymogens may be instigated by other enzymes or by a change in environment, for ex, pepsinogen (“ogen” at the end indicating zymogen status) is zymogen of pepsin and is activated by low pH
allosteric interactions
allosteric regulation is the modification of an enzyme’s configuration through the binding of an activator or inhibitor at a specific binding site on the enzyme
pepsin zymogen
the release of pepsin as a zymogen that is activated only by low pH ensures that pepsin only digests proteins where it is supposed to, in the stomach!
zymogen- pepsinogen
active enzyme- pepsin
function-digestive protease
allosteric regulation
- products that exert negative feedback inhibition do not resemble the substrate of the enzymes that they inhibit and do not bind to the active site
- instead they bind to the enzyme and cause a conformational change, which can be exerted by both allosteric inhibitors and allosteric activators
- not all are noncompetitve inhibitors which alter Km without affecting Vmax
allosteric enzymes
- meaning enzymes that have sites for allosteric regulation -do not exhibit typical kinetics because they normally have several binding sites for different inhibitors, activators and enzymes
- at low concentrations of substrate, small inc in concentration inc enzyme efficiency as well as reaction rate
- first substrate changes the shape of the enzyme, allowing other substrates to bind more easily, this is called positive cooperativity, opposite phenomenon negative cooperativity also occurs =cooperativity in presence of allosteric inhibitor is what gives the oxygen dissociation curve of hemoglobin its sigmoidal shape*
competitive inhibitors
compete with substrate by binding reversibly with non-covalent bonds to active site
- only type of reversible inhibitor that binds directly to active site rather than a different site on enzyme
- raise the apparent Km but do not change vmax***
- rate of reaction can be inc to the original, uninhibited Vmax by inc the concentration of substrate, since inc concentration of substrate is required to reach Vmax, an inc concentration is also required to reach 1/2 vmax
- Km raised showing lower affinity for enzyme for substrate
the ability to overcome inhibition….
by inc substrate concentration is the classic indication of a competitive inhibitor
uncompetitive inhibitors
-bind at site other than the active site -regulatory molecules can also bind to a site other than the active site and exert a positive feedback effect, rather than an inhibitory effect - do not bind to the enzyme until it has associated with the substrate to form the ES complex -once bound, substrate remains associated with the enzyme -the apparent affinity of the enzyme for the substrate increases, meaning that Km decreases -because this only affects enzymes that have already bound substrate, adding more substrate does not overcome the effect of inhibitor! -vmax is lowered because the substrate stays bound to the enzyme for a longer period of time
mixed inhibitors
-bind at site on enzyme other than active site, so they do not prevent the substrate from binding
-their names comes from the fact that they can bind to either the enzyme alone or the enzyme-substrate complex
- most have preference for one or the other which dictates the effect on Km and Vmax
- act like competitive inhibitors by binding primarily to the enzyme before the substrate is associated inc Km, just as competitive inhibitors do
- in contrast, mixed act more like uncompetitive inhibitors by preferring to bind to the enzyme-substrate complex lower Km
- all lower Vmax to some extent
noncompetitive inhibitors
special kind of mixed inhibitor
- bind just as readily to enzymes with a substrate as to those without, bind noncovalently to an enzyme at a spot other than the active site and change the conformation of the enzyme
- do not resemble the substrate, they commonly act on more than one type fo enzyme, they cannot be overcome by excess substrate, so they lower Vmax
- they do not however lower the enzyme’s affinity for the substrate because they bind to a site other than the active site, so Km remains the same
competitive inhibitors
- binding site= enzyme active site -inhibits binding of substrate
- effect on km= inc -effect on Vmax= no change
uncompetitive inhibitors
-binding site= Enzyme substrate complex -inhibits binding of substrate= NO -effect on Km= decrease -effect on Vmax= decrease
mixed inhibitors
- binding site= Enzyme substrate complex or enzyme
- inhibits binding of substrate= NO
- effect on Km= decrease or increase -effect on Vmax= decrease
noncompetitive inhibitors
- binding site= Enzyme substrate complex or enzyme
- inhibits binding of substrate= NO
- effect on Km= no change
- effect on Vmax= decrease
Line weaver burke
x intercept= 1/Km (correlates with enzyme substrate fit) y intercept = 1/vmax (max rate of catalysis) slope = km/vmax
Line weaver burke competitive
km inc, Vmax constant
Line weaver burke noncompetitive
Km stays the same/constant, vmax dec
differences btw ligase and lyase
-lyase particular type that catalyzes addition of one substate to the double bond of a second substrate is sometimes called synthase, ex. ATP synthase ligase enzymes require energy input from ATP or some other nucleotide*** sometimes called synthetases, so this is different than synthase* synthases do not require ATP to catalyze their reactions, while synthetases do
kinase
enzyme that phosphorlyates a molecule, often phosphoryaltes another enzyme in order to activate or deactivate it
can also be called a= Transferases catalyze transfer of a chemical group from one molecule (donor) to another (acceptor). Most of the time, the donor is a cofactor that is charged with the group about to be transferred. Examples include kinases and phosphorylases.
phosphatase
enzyme that dephosphorylates a molecule
enzymes
Enzyme is a catalyst
Usually a protein, but sometimes can be RNA (ribozyme) as well
Binds to one or more substrates
Reduces activation energy by stabilizing the substrate transition state
Lower activation energy yields increased reaction rate = catalysis
Lower activation energy may arise from enzyme helping orient reactants properly
enzymes 2
how they lower activation energy
- Lower activation energy may arise from enzyme helping orient reactants properly
- May arise from enzyme stabilizing transition state
- May arise from enzyme providing favorable environment for reactants (i.e., helping to stabilize charge)
- More transition states formed at given temperature means increased reaction rate
- Enzyme activity does not affect free energy (∆G) of the reaction or final equilibrium
- Enzyme is recycled at end of reaction
- Name of enzyme generally ends in –ase (kinase, polymerase, nuclease, ligase, protease)
enzymes mechanism and specificty
Induced fit model: slight change in enzyme shape upon substrate binding
Active site is where catalysis takes place
Active site is complementary to 3D shape of substrate, yielding specificity
In the active site, one or more amino acid residues are critical for catalysis
Active site residues interact with substrate and participate in reaction chemistry
Most enzymes are highly specific (e.g., act only on one of limited number of substrates)
Some enzymes are less specific (e.g., act on many substrates)
Which specific class of enzymes is primarily responsible for the release of free glycerol from stored triglycerides?
Q13 Blueprint exam 4
Lipases
-Lipases are the enzymes that digest lipids (fats). Most dietary fats originally exist in the form of triglycerides.
The diagram below depicts the six major classes of enzymes. Since lipases typically catalyze hydrolysis reactions, they are a subclass of the hydrolases.
B. Carboxylases= Carboxylases add carboxyl groups to their substrates.
C.
Phosphorylases= A phosphorylase is a transferase enzymes that moves phosphate groups between species. It does not relate to the digestion of fats.
D.
Kinases= A kinase is a transferase enzymes that moves phosphate groups between species. It does not relate to the digestion of fats.
ENZYME MECHANISM & SPECIFICITY
ex. induced fit vs lock and key
induced fit squeezes, there is a slight change in enzyme shape when substrate binds*
lock and key there is not a change in shape*
Induced fit model: slight change in enzyme shape upon substrate binding
Active site is where catalysis takes place
Active site is complementary to 3D shape of substrate, yielding specificity
In the active site, one or more amino acid residues are critical for catalysis, very important for the reaction* like some basic aa with positive charge, why certain kinds of mutations even if just change one amino acid, crucial can lose a lot of function for enzyme
Active site residues interact with substrate and participate in reaction chemistry
Most enzymes are highly specific (e.g., act only on one of limited number of substrates)
Some enzymes are less specific (e.g., act on many substrates)
Oxidoreductases 2
Oxidoreductases catalyze oxidation-reduction reactions where electrons are transferred. In metabolism, these electrons are usually in the form of hydride ions or hydrogen atoms. When a substrate is being oxidized, it is the hydrogen donor. Examples include reductases, oxidases, and dehydrogenases.
Lyases 2
Lyases catalyze reactions where functional groups are added to break bonds in molecules or they can be used to form new double bonds or rings by the removal of functional group(s). Decarboxylases are examples of lyases.
Lyases break covalent bonds using mechanisms besides hydrolysis
Isomerases 2
Isomerases catalyze reactions that transfer functional groups within a molecule so that a new isomer is formed to allow for structural or geometric changes within a molecule.
examples on graph
hydrolases 2
Hydrolases catalyze reactions that involve cleavage of a molecule using water (hydrolysis). This cases usually involves the transfer of functional groups to water. Hydrolases include amylases, proteases/peptidases, lipases, and phosphatases.
Hydrolases are responsible for the hydrolysis of bonds (proteases and lipase are examples)
Ligases 2
Ligases are used in catalysis where two substrates are stitched together (i.e., ligated) via the formation of C-C, C-S, C-N or C-O bonds while giving off a water (condensation) molecule.
Ligase joins two nucleic acid pieces
enzymes continued thermodynamics vs kinetics
do not change keq or delta G, can change kinetics but not thermodynamics
can make faster by stabilizing transition state, stabilizing charge, but they do not change the equilbirum position you end up with and they do not change delta G of the reaction
cofactors and coenzymes
Some enzymes require cofactors for action
Cofactors can be ions (e.g., Mg2+, Ca2+, Fe3+)
Cofactors that are organic, nonpeptide are coenzymes (e.g., coenzymeA, NADH, FADH2)
Most vitamins are coenzymes
Tightly bound cofactors are called prosthetic groups (e.g., heme in peroxidase)
Lots of kinases that require Mg2+, kinases transfer phosphate groups that are negatively charged have to have something stabilize negative charge, lots of enzymes have ot use Mg2+ positively charged, hangs out in active site, hangs there to donate a phosphate group, a ton of negative charge need some mechanism for stabilziing it**
prothestic group
cofactor tighly bound, not technically part of enzyme put attached like heme group*
temperature and enzyme activity
Enzymes have evolved peak activities at their physiological temperature
Optimal activity of human enzymes is near 37°C (body temperature)
Enzymes from other organisms may have other optimal temperatures
Example = Taq polymerase used in PCR is from thermophiles
Less enzyme activity at lower/higher suboptimal temperatures
At high temperatures, enzymes become denatured
Denaturation is caused by the loss of enzyme shape and structure, usually irreversible
pH and enzyme activity
Enzymes have optimal pH that reflects their catalytic environment
Most enzymes have peak ~ pH 7.2
Pepsin (protease in stomach) peak activity ~ pH 2
Lysosomal (hydrolytic) enzymes peak activity ~ pH 5
Chymotrypsin (pancreatic protease that functions in the intestine) peak activity ~ pH 8.5
Non-optimal pH reduces activity by altering enzyme charge and protonation
Adding or removing protons alters ionic bonds, H-bonds, and active site residues
Extreme pH causes irreversible denaturation
more basic environment more negatively charged, b/c base will go geting protons leave COOH- bases will deprotonate wherever they can, more positive enviroment acid constantly trying to stabilize more positive charges or stabilize engative charges, can be a very big deal if mess with how much positive or negative charge is in active site
zymogens 2
Some enzymes are synthesized as inactive pre-forms called zymogens
Zymogens are usually cleaved to give rise to active enzymes
Zymogen form prevents inappropriate enzyme activity (especially critical for proteases)
kd is like km
k2/k1 for Kd
- dissociation constant, doesn’t involve any formaiton of product
people say gives you a better estimate of affinity than Km does becuase it doesnt include anything to do with product formaiton** affinity talking about connection between enzyme and substrate, not whether ES complex goes on to form product***
if Kd is high** affinity is also low, and Kd is a slightly better proxy for affinity than Km but both are used and good to know both of them*
vmax 2
enzyme limit, where all active sites become saturated, if change how much enzyme you have can change vmax, premise is that you have limited amount of enzyme which limits your vmax* if Km high more falling apart less sticky, how fast going away of es complex versus how fast going toward which is opposite of affinity, why Km high thing falling apart low affinity
K2+ K3/ K1
Vmax = maximal reaction velocity = maximal turnover rate reached at saturating [S]
MM equation
when substrate equals Km, velocity equals 1/2 vmax
take reciprocal, graph it get straight line Lineweaver burkes so love it because means dealing with MM and can easily find out vmax, km x-intercept ,and y intercept in an easier way to figure out that original graph we looked at
k1 is rate constant for conversion of E + S to ES complex
k2 is rate constant for reverse reaction
k3 is rate constant for conversion of ES complex to E + P (also called kcat or turnover number)
reversible inhibition
- usually noncovalent bonds*** 3 categories compeitive, mixed (also includes noncompeitive), uncompetitive
- Reversible inhibition typically involves noncovalent interactions between inhibitor and enzyme
Reversible inhibition may be competitive, uncompetitive or mixed (including noncompetitive)
- Inhibition: may be reversible or irreversible
competivie inhibition 2
in presence of inhibitor Km goes up becuase affinity goes down, think about subtrate trying to bind ot enzyme but inhibitor gets in the way looks as though substrate and enzyme don’t like eachother as much don’t click just becuase interference happening so apparent Km inc so affinity APPEARS to go down just because inhibitor is interrupting the binding*
GRAPH= Y INTERCEPT STAYS THE SAME
Competitive inhibition: inhibitor mimics shape of substrate
Inhibitor binds to free enzyme
Inhibitor and substrate compete for active site
Can relieve competitive inhibition by adding more substrate
Maximal velocity Vmax is the same, but need more substrate to reach it
Apparent Km is increased because active site’s affinity for substrate is decreased
uncompetitive inhibition 2
- key difference inhibitor binds to ES complex, hovers and then once enzyme and substrate are together it gloms on, so weirdly the inhibitor stabilizes** the ES complex and prevents it from coming apart again** either in either direction, so prevents ES from forming product or going backwards to free enzyme plus substrate*
- jumping to apparent Km, weird thing is looks affinity is very high seems like substate and enzyme really love eachother totlly stickign togethe rbut just becuase inhibitor is pouring glue* all over ES complex* it freezes the ES complex in place, sticks it together in a way that inhibits reaction but makes it see as if es are super super sticky and high affinity, meaning looks like Km is very low Km goes down**
- vmax decreases because no way to flood system with substrate and overcome the effects of the inhibitor** Km goes down, Vmax also goes down!! but go down proportionally to eachother, so that meanas that the slope of the graph with and without inhibitor is the same** so that means if look at the graph get parallel lines all the same slope whether have a little bit of inhibitor, a lot of inhibitor or what not** when look at graph its the slope that stays the same so you have parallel lines*
mixed inhibition
- the system inhibition will be sort of liek competitive and sort of like uncompetitve, vmax always goes down becuase not 100% competitive, but impact on Km can go eitehr way
- competitive inhibition Km goes up adn uncompetitive inhibition Km goes down, so can be either/or depending on the specifics enzymes and inhibitor
- recognize this because doesnt look like anything is staying constant*** yintercepts are diff, slope diff, x intercept diff, lines cross at some random place not on an axis**
noncompetitive inhibition 2
- mixed is blend of competitive/uncompetitive, competitive makes Km go higher, uncompetitive makes Km go lower, so somewhere in there balance btw these two and the Km will not change
- This is a tiny sliver of mixed inhibition cases* but the ones where the Km stays exactly the same, those are called noncompetitve inhibition*** when Km is hte same, then the xintercept is the same*
- xintercept on graph ex is -1/Km so what you are looking for is lines radiating out from X axis there*
Q11
what the cofactor is doing is stabilizing phsopahte groups**, stabilizing negative charges etc, work in active site*** allosteric site are elsewhere, and they help binding of susbtrate wouldn’t prevent it they are facilitating reaction!
allosteric sites:
can be involvd in enzyme activation, enzyme cooperativity, and enzyme inhibition
can have diff molecules bind to allosteric site activators or inhibitors, engative allosteric regulation inhibiting reaction*
uncompetitive inhibition 3
Uncompetitive inhibition
Inhibitor binds to ES complex, stabilizes ES complex in transition state
Inhibitor does not bind at active site
Vmax decreases because inhibitor slows enzyme activity. Less S → P
Effect of enzyme cannot be overcome by adding more S
Km decreases because active site’s affinity for S is increased (enzyme can’t release S)
enzymes details
Enzyme is a catalyst
Usually a protein, but sometimes can be RNA (ribozyme) as well
Binds to one or more substrates
Reduces activation energy by stabilizing the substrate transition state
Lower activation energy yields increased reaction rate = catalysis
Lower activation energy may arise from enzyme helping orient reactants properly
May arise from enzyme stabilizing transition state
May arise from enzyme providing favorable environment for reactants (i.e., helping to stabilize charge)
More transition states formed at given temperature means increased reaction rate
Enzyme activity does not affect free energy (∆G) of the reaction or final equilibrium
Enzyme is recycled at end of reaction
Name of enzyme generally ends in –ase (kinase, polymerase, nuclease, ligase, protease)
mixed inhibition 3
Mixed Inhibition
Inhibitor binds to free enzyme AND to enzyme-substrate complex
Inhibitor does not bind to the active site
Exhibits properties of both competitive and uncompetitive inhibition
Substrate binding affects inhibitor binding and inhibitor binding affects substrate binding
Vmax decreases because enzyme activity is slowed by inhibitor (less effective S → P)
Can be partly but not fully overcome with increased S
Effects on Km depend on inhibitor binding:
Km increases if inhibitor binds preferentially to E alone (harder for S to bind)
Km decreases if inhibitor binds preferentially to ES complex (S cannot unbind)
noncompetitive inhibition 3
Noncompetitive Inhibition
Special case of mixed inhibition
Vmax decreased because enzyme activity slowed by inhibitor
Cannot be overcome by adding more S
Km is unchanged because the active site’s affinity for substrate is unchanged
In practice very few examples of pure non-competitive inhibition are known
irreversible inhibition
Irreversible inhibition: inhibitor inactivates enzyme permanently
Inhibitor reacts with and destroys active site or other critical residues Tight or covalent binding to enzyme (also called suicide inhibitor)
enzymes with 2 or more substrates
Numerous reactions in the body involve an enzyme interacting with two different substrates
Enzyme and two substrates bound together → ternary complex
Ordered binding: one substrate must bind before the other. When both have bound → ternary complex
Random binding: either of the two substrates can bind first. When both have bound → ternary complex
Ping pong mechanism: no ternary complex
In ping pong mechanism, one substrate forms ES complex and first product is produced. Enzyme is still in a modified form.
Second substrate forms ES complex, and second product is produced. Enzyme is regenerated in original form.
Ping pong mechanism is also called double displacement
ordering binding
Ordered binding: one substrate must bind before the other. When both have bound → ternary complex
random binding
Random binding: either of the two substrates can bind first. When both have bound → ternary complex
ping pong mechanism
Ping pong mechanism: no ternary complex
In ping pong mechanism, one substrate forms ES complex and first product is produced. Enzyme is still in a modified form.
Second substrate forms ES complex, and second product is produced. Enzyme is regenerated in original form.
Ping pong mechanism is also called double displacement
allosteric enzymes 2
Allosteric enzymes have more than one conformation (shape)
Different conformations have different enzymatic activity
An allosteric regulator binds to enzyme and triggers conformational change
Allosteric regulation can be positive (increase activity) or negative (decrease activity)
allosteric activation
Allosteric activation: a chemical species binds to enzyme usually at allosteric site
Increases the rate of reaction
Does not necessarily involve cooperativity between subunits
cooperativity
Cooperativity = substrate binding to one site changes affinity of other sites for substrate
Typically involves enzymes with multiple subunits
Can be positive (i.e., substrate binding increases activity of other subunits) or negative
A positively cooperative enzyme has a more sigmoidal curve for enzyme activity
Cooperative = “Hill coefficient > 1”
nuclease
Nuclease cuts nucleic acid
DNAse is nuclease that digests DNA
RNAse is nuclease that digests RNA
exonuclease
Exonuclease cuts at ends of nucleic acid
endonuclease
Endonuclease cuts within nucleic acid
polymerase
Polymerase synthesizes (polymerizes) nucleic acids
protease
Protease cuts protein
kinase
Kinase adds a phosphate (is a type of transferase)
phosphatase
Phosphatase removes a phosphate
mutase
Mutase moves a phosphate group within molecule
dehydrogenase
Dehydrogenase catalyzes redox reaction. Usually involves NAD+ or FAD
glycosylase
Glycosylase removes sugars
carboxlyase
Carboxylase usually transfers CO2, adding a carboxy group to the substrate. Often requires biotin as a cofactor.
biological regulation of enzyme activity
Can change rate of enzyme synthesis (e.g., transcription or translation)
Enzyme can be activated by processing (e.g., zymogen)
Enzyme can be regulated by post-translational modification (e.g., phosphorylation)
Feedback inhibition: enzyme is inhibited by a product of a reaction sequence
Product binds to enzyme and inhibits activity
Feedback inhibition is commonly used in biochemical pathways
Example: End product D inhibits enzyme 1 in rxn sequence A (1)→ B (2)→ C (3) → D
end product D can inhibit enzyme 1, (enzyme)
Consider Enzyme B at a point below Vmax. As substrate concentration is increased and an allosteric activator is added, how will reaction rate and Vmax be affected?
A.Both Vmax and the reaction rate will remain constant.
B.Both Vmax and the reaction rate will increase.
C.Vmax will remain constant, while the reaction rate will increase.
D.Vmax will increase, while the reaction rate will remain constant.
C.
Vmax will remain constant, while the reaction rate will increase.
CORRECT Vmax is not dependent on substrate concentration, and it does not change upon addition of an activator. In fact, Vmax would increase only if more enzyme were added. However, increased substrate concentration does raise the reaction rate, as is shown by the graph; exposure to an activator should have the same result.
Chymotrypsinogen
Chymotrypsinogen= zymogen
active enzyme= Chymotrypsin
function= digestive protease
tyrysinogen
zymogen= tyrysinogen
active enzyme= trypsin
function= digestive protease
Procarboxypeptidase
zymogen= Procarboxypeptidase
active enzyme= Carboxypeptidase
function= digestive protease
Proelastase
zymogen= Proelastase
active enzyme= Elastase
function= protease
Prothrombin
zymogen= Prothrombin
active enzyme= thrombin
function= protease for clotting
enzyme kinetics
Key aspect of enzyme catalyzed reaction is saturation kinetics
Once enzyme is saturated, additional substrate cannot accelerate the reaction
S = substrate concentration
V = reaction velocity
Vmax = maximal reaction velocity = maximal turnover rate reached at saturating [S]
E = free enzyme
ES = enzyme-substrate complex
At low [S], reaction rate increases linearly with increasing [S]
At high [S], reaction rate reaches a maximum (enzymes are saturated)
Michaelis-Menten kinetics: graph of v0 versus S is hyperbolic
Km 2
Km = the Michaelis constant
Km = an approx. measure of the apparent substrate binding affinity for the enzyme
V0 = Km X [S]/ Km + [S]
When [S] = Km, v = 1⁄2 Vmax
High Km → low affinity of substrate for enzyme (more S needed for 1⁄2 Vmax)
Low Km → high affinity of substrate for enzyme (less S needed for 1⁄2 Vmax)
Kd
Kd = dissociation constant = k2/k1
Most accurate measure of affinity of enzyme for substrate
Ka = association constant = 1/Kd
Vmax = kcat × [E]
Doubling the concentration of enzyme will double the Vmax
Catalytic efficiency = kcat/Km
Lineweaver-Burke:
take reciprocal of both sides of Michaelis-Menten equation:
Plot 1/[S] on x-axis; 1/v0 on y-axis
If enzyme obeys Michaelis-Menten kinetics, Lineweaver-Burke → straight line Slope = Km/Vmax
Y-intercept = 1/Vmax
