Transcript Catabolism vs Anabolism

BIBC 102
Metabolic Biochemistry!
Lectures #1 and #2!!
• Hello. I am Tommy Wootton.
• Feel free to shut me up any time for
questions!
Catabolism vs Anabolism
To break down
(AKA Cut-abolism!)
To build up
Anabolic steroids!
know the 20 amino acids by
name, three letter code, one
letter code, and be able to
recognize (as opposed to
draw) the structures.
Amino
acids:
The
Molecular
Tool Box
(G)
(L)
(D)
(M)
(I)
(S)
(P)
(T)
(N)
(C)
(Q)
(E)
(K)
fig3-5
(V)
(A)
(R)
(H)
(F)
(Y)
(W)
Amino Acid/Peptide Bond
Protein Structure
Primary Structure - AA sequence
Secondary Structure - 3D structures! ( helix,  sheets)
Tertiary Structure - Interactions between these
secondary structures
Quaternary Structure - Multiple subunit interactions
So, lets use a word analogy…
“This guy SUCKS!”
-Hopefully NOT any of you
Primary? --> letters
Secondary? --> words
Tertiary? --> sentence
Quaternary? --> multiple sentences interacting to
achieve satisfactory expression of distaste
Energy map of a reaction
fig6-2
DG‡ is the activation energy
Activation energy and reaction rate
fig 6-3
4 Main Enzymatic Catalytic Mechanisms
Entropy Reduction
Acid Base Catalysis
Metal Ion Catalysis
Covalent Intermediates
Entropy Reduction
Bringing the pieces together! Forcing them to interact.
Think of Randy’s
analogy with 2 cats.
Or a cat and a dog.
If they’ve got plenty of
space, they’re not
going to interact. But
if they’re brought
together…
Acid Base Catalysis
Providing or picking up protons to allow for
more efficient reaction catalysis.
Happens in Chymotrypsin!
H+ provided by Ser195 OH
group!
Metal Ion Catalysis
Involves the stabilization of intermediate structures
through ionic bonding from metal ion cofactors like Zn2+.
Haven’t talked about it much, but some enzymes do it.
Covalent Intermediates
When a substrate forms a covalent bond with the enzyme
to form an intermediate!
Remember TPP (Thiamine Pyrophosphate) or the
dihydrolipoyl arm from Pyruvate Dehydrogenase
Complex?!
Sample Problem!
free
energy
reaction
Ligand Binding
The dissociation constant (Kd)
S + E (Separate)  SE (bound together!)
Kd
[S][E]
= ____
[SE]
SO…
If Kd is BIG, S and E tend to be separate
(HIGH dissociation, get it?)
If Kd is little, S and E will tend to be bound
So back to ligand enzyme binding equation…
If Kd = 0, LB = B (Saturation! Every emzyme is bound)
BUT, if Kd is big (=enzyme and ligand prefer to be separate),
LB becomes a smaller proportion of B
What if Kd = L? Hmmm…
Now something cool…
If Kd = L, then LB = 1/2 [B]
So it can tell you (and let you compare!) the affinities of an
enzyme to a ligand in different conditions…
Related to the Michaelis-Menton Equation?
Yep! Pretty similar. Except Michaelis-Menton
represents enzymatic rates, NOT ligand binding
Vo = Vmax
(
)
[S]
______
[S] + Km
LB = B
(
)
[L]
_____
[L] + Kd
Same general equation! So, same general shape…
So really, just a change in axes labeling!
- Km = efficiency of RXN
- Kd = efficiency of binding
- Gives info about enzyme activity
-Gives info about ligand
binding
What about a cooperative
enzyme?
Due to quaternary
structure. Multiple
subunits and binding
afinity increases as
each is bound.
Makes Vo increase as
[S] increases (until
you approach Vmax
and saturation.
Ex. Heme vs.
Myoglobin
So how fast can an enzyme work?
Vmax = Kcat [E]total
Whats Kcat?
The rate at which an individual enzyme
can undergo catalytic function. Essentially it’s how quick
each individual enzyme molecule can do it’s work.
Sample Problem!
M is half saturated when S equals the value of Km… 5 uM
S/(S + Km) = 20/25. So 0.8, or 4/5, or even 80% are acceptable
L is half saturated when S equals the value of Km… 20 uM
The ratio is 1. Since the kcats are the same, the maximal rates
(for a given amount of enzyme) are the same.
Three Types of Inhibitors!
1. Competitive Inhibitor
2. Uncompetitive Inhibitor
3.“Suicide”
Inhibitor
Competitive Inhibitors
*not quite like this!
The dumb@$$ in this
picture would be a
dysfunctional ligand…
BUT it’s about the
concept. They take up
space AT the active
site.
What does this look like
molecularly?
One more view…
Uncompetitive Inhibitors
You’re almost there. At the computer (AKA active site).
Ready to go, but someone (inhibitor) is bugging ya.
An uncompetitive
inhibitor binds away
from the active site.
Doesn’t compete for
the active site. Not
like in this pic.
Looks like?
Another view…
“Suicide” Inhibitor
Literally takes the computer
(enzyme) out of commission.
Covalently binds to the
enzyme and breaks it down
so it doesn’t work anymore.
-Lowers Vmax.
-No effect on Kcat
:(
Allosteric regulation
Allo = other, steric = site SO, regulation through
binding at a site OTHER than the active site.
Can be inhibitors OR activators. Often the regulator is
a product or substrate (allows for negative feedback!)
Chymotrypsin
A serine protease! Cuts up proteins! Hydrolyzes its
substrates (W, Y, F, L) at the carboxy terminal of the
peptide.
W=Tryptophan, Y=Tyrosine, F=Phenylalanine,
L=Leucine…
The Catalytic Triad = Asp102, His57, and Ser195.
Asp102 stabilizes the positive charge of His57 when it
picks up proton from Ser195 after nucleophilic attack.
The Catalytic Triad = Asp102, His57, and Ser195. Hydroxy
group of Ser 195 attacks. Proton transferred to His57.
Asp102 stabilizes the positive charge of His57 when it picks
up proton from Ser195 after nucleophilic attack.
Lineweaver-Burk Plots!
A couple things to remember:
-Axes are reciprocals
-Tells you Vmax, Km
Know where they are!
-Be able to think about it.
Talk through it. Shouldn’t
just look like random lines!
Competitive Inhibitors!
As [I] increases,
Km increases, but
Vmax stays the
same!
This is because
you will need more
S to reach 50%
saturation (to displace
I), but the max rate
can theoretically still
be reached! Just
takes more S.
Uncompetitive Inhibitors
Here, Vmax decreases
as [I] increases. Also,
Km decreases!
This is because I binds
to an allosteric site and
while bound, makes
E inactive.
Sample Problem!
1/[S]
1/Vo
-1/Km
1/Vmax
When Michaelis-Menton enzymes are plotted in this manner, the
resulting data is a straight line, making it easier to see when an
enzyme behaves this way and easier to discern what kind of
inhibitor is involved in an experiment.