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Enzymes as Proteins. Enzymes Kinetics. Regulation of Enzyme Activity


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Do enzymes change the equilibrium of a rxn?
No... only rate (or how fast the rxn reaches equilibrium)by lower activation energy
In E + S <-> ES... name the reactants and products
E = enzyme
S = substrate
ES = Michaelis complex (reversible)
In A --> P, how do you determine the rxn velocity... and describe its curve
v = k [A]
a straight line
In an ezyme catalyzed rxn, what eventually happen when your substrate concentration saturates the enzymes?
V-max... at ES complex formation
At low [substrate], what would you expect?
velocity proportional to [S]... a straight line
So, with respect to the work of the enzyme, what is happening at v-max?
it's working as fast as it can, when a substrate leaves the binding site another enters.
a. When is v virtually independent of [S]?
b. and what is the rate order at that point?

a. At high concentrations
b. zero order
what portion of the total enzyme do active sites take up of the surface?
What is going on in the active site of an enzyme?
residues directly participate in the making and breaking of bonds.
In terms of the active site, what is important the sequence or the folding of the residues?
most important is the folding which is actually a result of the sequence.
How does the enzyme recognize the substrate?
a. electrostatic force
b. hydrogen bonding
c. van der Waals interactions d. hydrogen bonds
e. various hydrophobic interactions in which water molecules are stripped off the substrate.
What makes the enzyme and the substrate come together?
Hydrophobic effects
What role do the enzymes play in attracting the substrate?
the crevices are hydrophobic, which excludes water and enhances catalysis. (like an SN2 rxn where a nucleophile and an electrophile can come together.)
Was Emil Fischer right about the lock and key hypothesis?
What is Koshland's Induced fit hypothesis?
After binding to the substrate, the enzyme undergoes a conformational change.
What effect does an enzyme have that causes the lowering of the activation energy and thus speeds up the rxn?
They stabalize the transition state.
What is a Kcat or k2 number?
the number of substrate molecules converted to product by a single enzyme per unit time.
What is the Michaelis-Menten equation?
v = k2[ET][S]÷km+[S], where
v-max = k2[ET]
k2=kcat conversion per enzyme
[S} = [substrate]
km = a given substrate constant... thus,
v= Vmax[S]÷ Km + [S]
What happens when the substrate concentration is high... or [S]≥ Km?
Ignore Km, thus, [S] cancels and...
V = Vmax
What happens when the [S]< Km?
v = (Vmax/Km)[S]
a. now km is much more important
b. and the substrate demoninator is ignored.
what it the velocity of a rxn when km=[S]?
a. What happen if you're near your Km and you double or half your [S]?
b. How is this relevent to cells?
a. It makes a big difference
b. if the [S] is near its km then varying it's [S] can have a big effect on the rate of product formation.
Since V-max is never reached, how do you determine Km, if you don't know Km you don't know V-max?
a. Take the inverse of velocity / the inverse of the [S], then
b. take two points and draw a staight line, then
c. y-int = 1/V=max
d. x-int = -1/Km
What is acetylpepstatin?
An HIV protease inhibitor
What effect does a small KI (for an inhibitor) have on the rxn rate?
small KI means more EI (inhibitor in the enzyme) and less enzyme is available for the substate.
ON the Lineweaver-Burk plot, what does a sleeper slope when a competitive inhibitor is added?
steeper slope, slower rxn... not as much enzyme available for substrate.
How will a competitive inhibitor effect the V-max?
it will have no effect... 1/V-max is always at the Y-int... you just have to add more substrate to out compete the competitive inhibitor to achieve V-max.
Which 3 amino acids undergo phosphorylation
serine, threonine, tyrosine
a. Is an enzyme activated by a protein kinase or a protein phosphatase (in the presense of ATP or water)?
b. is this reversible?
phoshorylation or dephosphorylation can activate or deactivate.

it is reversible
a. In protein kinase phosphorylation, what is the phosphoryl donor?
b. In protein phosphatase dephosphorylation, what is the phosphoryl acceptor?
a. ATP
b. H2O
Is phosphorylation permanent?
No, these rxns are reversible
Is prenylation permanent?
What is prenylation?
It is the adding of 5-C aliphatic moiety (very lipophilic) that anchor proteins to the membrane... (Farnesyl and Geranylgeranyl)
What is the role of ras
The role of Ras is to send a growth-promoting signal through a MAP kinase cascade
What two proteins phosphorylate and dephosphorylate ras? and which is activating?
GEF phosphorylates - activates
GAP dephosphorylates
Why is it that inhibiting the prenylation of ras a possible cancer therapy?
Prenylation traps ras at the membrane where it activates the kinase cascade... freeing it will inhibit its role.
what is an allosteric enzyme?
Allosteric enzymes are enzymes whose activity is modulated by small molecules that reversibly bind to a site other than the active site... changing the enzymes conformation.
Are allosteric enzymes activating or inhibiting?
Either one
T/F: all enzymes exhibit Michaelis-Menten kinetics.
False, allosteric enzymes do not.
which type of enzyme gives a hyperbolic curve and which a sigmoid curve?
a. allosteri enzyme = sigmoid
b. non-allosteric enzyme = hyperbolic.
Which type of enzyme is often inhibited by the end product?
allosteric enzyme.

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