BCOR 011
Lecture 12
9/28/2005
ENZYMES
Last time…
-G
reaction “can” go
spontaneous
But when will it go?
And at what rate?
Thermodynamics
Whether a reaction will occur
Kinetics
WHEN a reaction will occur
What governs WHEN a reaction will occur?
The tower of blocks falling is favorable
but when will it happen?
Oxidation of carbohydrate polymers (starch)
to carbon dioxide and water is favorable
but when will it happen?
Gasoline burning to carbon dioxide and water
is favorable
but when will it happen ?
For a Reaction to occur need to Destabilize Existing State
to INPUT ENERGY
Destabilization
energy input
“Activation
Energy”
Potential
net
usable
energy
Now
Potential
net
usable
energy
In Transition
Need to INPUT ENERGY to Destabilize Existing State
Regain
Activation
Energy
Invested
net
usable
energy
released
Potential
net
usable
energy
In Transition
After
What does activation energy represent?
For a Reaction to Occur…
- reactants must find each other,
- meet in proper orientation
- and hit with sufficient force
Productive
Collision
Many
Non-productive
Collisions
Needs of Typical
chemical reactions
- need large number of molecular collisions
- need collide violently enough to
break pre-existing bonds (not bounce)
- need high concentration to find each
other at significant rate
HEAT !
The energy profile for an exergonic reaction
A
B
C
D
Free energy
Transition state
A
B
C
D
EA
Reactants
A
B
∆G < O
C
D
Products
Progress of the reaction
Figure 8.14
Temp 1
Temp 2
EA
Molecules with
sufficient
Energy (~40%)
Molecules with
sufficient
Energy (<5%)
ENZYMES make reactions
easier to occur at
reasonable temperature
by
LOWERING the
ACTIVATION ENERGY
EA
of the reaction
Activation Energy
Energy necessary to overcome the status quo
EA
G
EA
G
“ease” of
initiating reaction
Thermodynamic
“favorablility”
CATALYSTS:
promote a specific reaction
But are NOT consumed in the process
Key concepts:
Promotes
- does not alter what would normally
occur thermodynamically
Specificity
- promotes only one reaction, only
between specific reactants to give specific products
Reusable
- regenerated in the process
ENZYMES are biological CATALYSTS
- usually PROTEINS
- sometimes RNA or
RNA/protein complexes
Enzymes work as catalysts
by providing an easy path to the same point
Hard path
Easy path
HOW?
How do Enzymes do it?
1. Enzymes have BINDING AFFINITY
for their reactants = Substrates
Brings substrates in close proximity: conc
Stabilized Interactions
Charged
Nonpolar
Polar
Have a very Specific 3-D Shape
With a Specific Arrangement
of Functional Groups
Flexible
OH
HO
HO
+
Enzymes act as a Specific Platform
ENZYMES:
Bind ONLY specific things
HO
OH
Bind them ONLY in a
Specific 3-D Orientation
OH
OH
HO
-
HO
+
SPECIFICITY is the Key to Enzyme Action
2. Enzymes ORIENT Substrates
always in productive orientation
Productive
Collision
Many
Non-productive
Collisions
ONLY Productive
Collisions
With just a little nudge, can’t help but react
HO
OH
OH
OH
HO
+
HO
3. Enzymes cause BOND STRAIN
- destabilize existing bonds
“nutcracker effect”
3a. Physical Strain
3b.Chemical Strain
The active site
– Is the region on the enzyme where the
substrate binds
Substate
Active site
Enzyme
Figure 8.16
(a)
Induced fit of a substrate
Enzyme- substrate
complex
Figure 8.16
(b)
Enzyme-substrate interactions
Fischer:
Lock & key
Koshland:
Induced fit
3a. Physical bond strain
Draw an quarter - an anvil
• The catalytic cycle of an enzyme
1 Substrates enter active site; enzyme
changes shape so its active site
embraces the substrates (induced fit).
Substrates
Enzyme-substrate
complex
6 Active site
Is available for
two new substrate
Mole.
Enzyme
5 Products are
Released.
Figure 8.17
Products
2 Substrates held in
active site by weak
interactions, such as
hydrogen bonds and
ionic bonds.
3 Active site (and R groups of
its amino acids) can lower EA
and speed up a reaction by
• acting as a template for
substrate orientation,
• stressing the substrates
and stabilizing the
transition state,
• providing a favorable
microenvironment,
• participating directly in the
catalytic reaction.
4 Substrates are
Converted into
Products.
3b. Chemical Bond Strain
tease the bond to fall apart
Chemical Bond Strain
Stabilize a
Fictitious
state
Cofactors
Non-polypeptide things at the active site
that help enzymes do their job
• Cofactors
– Are nonprotein enzyme helpers, eg Zn++
• Coenzymes
– Are organic cofactors
4. Enzymes “partake” in reactions
but are not consumed in them
Converts MANY “A’s” into “B’s”
H+
H+
OH-
Partakes: but start and end with the same enzyme config
Lysozyme
Lysozyme: kills bacteria
Works at pH 4-5 Why?
SUMMARY
Enzymes:
1. Bring reactants (substrates) in close proximity
2. Align substrates in proper orientation
3. Can act as a Lever: a press or an anvil
small shape change translates to large force
4. Release products when reaction done
rebind more substrates
5. Many small steps, each easily achieved
rather than one huge leap
Enzymes carry out reactions in a series of
small steps rather than one energetic event
No Problem
Dude
You expect
me to
JUMP this?
Reaction rates:
Example: H2O2-> H2O +O2
uncatalyzed –months
Fe+++ 30,000x faster
Catalase 100,000,000 x faster
Enzyme kinetics- kinetikos – moving
An enzyme catalyzed rxn
Can be “saturated”
Vmax
maximum velocity
Rate
or
velocity
1/2 Vmax
# made
per min
Km
Substrate Conc
“substrate affinity”
The lower the Km the better
the enzyme recognizes substrate
“finds it at low conc”
“mpg”
The higher the Vmax the more
substrate an enzyme can
process per min
(if substrate around)
“top speed”
Things that affect protein structure
often affect enzyme activity
temperature
0
20
40
60
80
100
ºC
pH
0
1
2
3
4
5
pH
6
7
8
9
10
Enzyme regulation:
Activity controlled
Continually adjusted
Principal Ways of Regulating Enzymes
Competitive Inhibition
Allosteric Inhibition
Covalent Modification (phosphorylation)
Competitive
Inhibitors:
bind to active site
“unproductively”
and block
true substrates’
access
HO
S2
S1
OH
OH
-
I
HO
OH
OH
HO
HO
S & I bind to same site
+
Competitive inhibition
Allosteric Inhibitors
“other” “site”
Distorts the conformation
of the enzyme
Negative
allosteric
regulator
Allosteric inhibition
Positive allosteric regulators
Helps enzyme work better
promotes/stabilizes an “active” conformation
Allosteric activation
Allosteric regulators change the shape
conformation of the enzyme
Allosteric enyzme
with four subunits
Regulatory
site (one
of four)
Active site
(one of four)
Activator
Active form
Stabilized active form
Oscillation
Allosteric activater
stabilizes active form
NonInactive form Inhibitor
functional
active
site
Figure 8.20
Allosteric activater
stabilizes active from
Stabilized inactive
form
(a) Allosteric activators and inhibitors. In the cell, activators and inhibitors
dissociate when at low concentrations. The enzyme can then oscillate again.
A frequent regulatory modification
Phosphorylation of enzymes
Phosphorylase kinase
inactive
+ P
active
Summary
1.enzymes are catalysts
2.Lower activation energy EA
3.Mechanism of action …
4.Enzyme kinetics- Vmax, Km
5.Regulation of enzyme activity
- competitive, allosteric
phosphorylation