Ch 8. Energy, Chemical
Reactions, Enzymes, and
Metabolism
Figure 5.00a
Figure 5.00b
Figure 5.00c
As the potential energy of a system is released (here
converted to kinetic energy) the system becomes more stable
and the released energy is available to create change (do
“work”) within the system. Similar energy conversions govern
cellular metabolism.
The Laws of thermodynamics govern energy flow in
living systems the same as in the non-living world.
Energy can’t be created nor destroyed, it only
changes form; AND when it does there is always less
usefull (ie. “free” ) energy left afterward
In living cells, some of the energy liberated in a “breakdown” or catabolic
reaction can be saved in the form of ATP, but much is still lost as heat.
This slide was included in the Ch 7 presentation; lets
look at what’s going on here in a bit more detail
ATP structure
Cells consume ATP at a phenomenal rate so it must constantly be
regenerated using the energy supplied from the environment as
sunlight or chemical bond energy
Catabolic (“breakdown”) reactions liberate chemical
bond energy, some of which is captured and transferred
to ATP via linking ADP and P.
Anabolic (“build-up” or synthesis) reactions require an
external source of chemical bond energy, which is
usually supplied by the hydrolysis of ATP to ADP and P.
Some reactions are “spontaneous” - they go with
the flow, while other reactions are not.
A spontaneous reaction involves an energy change that is in
keeping with the second law of thermodynamics which means
that as the reaction proceeds energy is released and the
system contains less high quality energy after the reaction than
before.
A nonspontaneous reaction works has the opposite properties
and so requires a high quality energy input.
Both are possible in cells, and both can be catalyzed by
enzymes.
All chemical reactions involve some sort of free (ie. “usefull”) energy
change G. Free energy is either released (left) or an input required
(right) as the reaction progresses.
If more is released, then the products contain less than the reactants (red rxn
on left) and the free energy change G is negative (-). Such rxns, in effect,
work in accordance with the second law of thermodynamics - they “go with the
flow” so to speak, and are referred to as being “spontaneous.”
The (Gibbs) Free energy change that
accompanies a reaction is a
composite of the change in chemical
bond energy or enthalpy, H, and the
change in the product of temperature
(in oK) times entropy, TS that a
reaction produces.
G = H - TS
Josiah Willard Gibbs was a bachelor engineer who received his Ph.D. and did
all of his subsequent work at Yale. He was reclusive genius who founded the
field of chemical thermodynamics in he late 1800’s. The effort involved in
comprehending his work is still driving students (not to mention professors)
crazy to this very day. Gibb’s work received somewhat limited acclaim until
1923 when Lewis and Randall publishedThermodynamics and the Free
Energy of Chemical Substances, which introduced the methods of Gibbs to
chemists world-wide. (Wikipedia)
To confuse the issue further, we can express G in terms of the
concentration of reactants and products at the equilibrium point.
Where Keq for the reaction A+B -> C +D or
is defined as:
An oversimplified (but less mind-bending for the symbolically
challenged) way to look at this is that the equilibrium
constant depends on the relative concentrations of the
reactants (A and B) and products (C and D) at the point in
the reaction where neither is changing any further.
So a reaction’s Keq is calculated from measurable
quantities, and therefore the subsequent calculation of G
can be done based on actual measurements for any set of
conditions.
A key point here is that the Keq and G for a reaction is
dependant only on the chemical nature of the reactants
and products, their concentrations, and the conditions
under which the reaction is carried out. The speed at
which the reaction occurs is not relevant (it’s independent).
So, what’s all this have to do
with enzymes?
Hold that thought and consider the
following analogy based on jumping
beans (its not perfect, but few
analogies are).
Jumping beans are seeds of a Euphorb shrub, Sebastiania palmeri or S.
pavoniana that have become infested with the larva of a moth, Cydia deshaisiana,
these are found in the Mexican desert. The larva consume the seed contents and
persist within the seeds even after they’ve fallen from the plant. Temperature
changes cause the larvae to contract violently causing the whole seed to “jump.”
The beans will often “jump” at random times
with random orientations (in any direction) and
energies.
So “beans” in a box will jump with an
occasional jump being energetic (high) enough
and oriented such that the bean clears the box
walls
OK, so lets let the beans in the left
hand box represent reactants and
those in the right hand box represent
products. I know, you can argue that,
unlike real reactants and products,
the beans are still the same
regardless of which box they’re in,
but let’s ignore that.
Given enough time, some beans would
manage to jump the barrier (wall)
separating the two compartments.
That is, “reactants” would become
“products”.
As pictured, the “product” beans are unlikely to
be able to jump back and become reactants
again due to the difference in the depth of the
compartments.
H
This is analogous to a change (a decrease) in
H between reactants and products that would
be expected for a (exothermic) “spontaneous”
reaction where G is negative and large (free
energy is given off)
In some cases, both H and S ( both the height and width of the boxes)
differ. A given # of molecules occupying a larger space = an increase in
disorganization or entropy ( an increase in S). Such a reaction in which H
decreases and S increases, would be strongly exergonic and spontaneous.
The combustion of paper (combining O2 and cellulose or linked glucose) to
CO2 and H2O for example.
S
So what’s all of this got to do with enzymes? The short
answer to this point is, “nothing.”
The G, H, and S of any reaction, as stated before, are
functions of the chemical nature of the reactants and products
and/or the conditions under which the reaction is carried out.
Regardless of whether an enzyme is involved or not.
An enzyme will make it possible for an energetically feasible
reaction to occur faster, it won’t make a non-feasable
reaction possible.
Enzymes accelerate reactions; they can’t affect the extent to
which a reaction goes to completion (the Keq) or theG.
This acceleration occurs via the lowering of the activation
energy barrier.
Enzymes are biological catalysts that allow the
chemical reactions of life to occur much faster than
they could otherwise (with a much lower energy
input).
• They are mostly catalytic proteins.
• Very specific (have specific substrates)
• Not consumed
• Activity can be modulated (controlled)
•They don’t alter the nature of the energy change that a given
reaction entails, they merely make “possible” reactions go
faster.
For the combustion of glucose
Without an enzyme, the large activation energy requirement must be
supplied by the application of an external source of heat. In cells,
glucose is “combusted” in many small steps the combined G of
which is the same as if it had all been accomplished in one step as
above.
The key to reducing the needed activation energy is the holding of
the reactants in the active site in a manner such that less
activation energy is needed to break old bonds or make new
ones.
Another version:
Reactions that are non-spontaneous, such as synthesis reactions
(more chemical bonds and order are being created), require an
energy input to provide the necessary + G. This is usually
accomplished by linking such reactions with the hydrolysis of ATP;
the high energy P so produced is first linked to a substrate(s) to
provide the needed + G.
To synthesize the amino acid glutamine from glutamic acid the glutamate is
first phosphorylated via ATP hydrolysis.
Remember, Enzymes are:
Highly substrate-specific.
Affected by the relative concentrations of enzyme and
substrate to the point where the reaction rate is saturable.
Subject to competitive and non-competitive inhibitors.
Subject to (affected by) the physical (temperature) and
chemical (pH, [salt]) conditions under which they operate.
Most often operate as part of a metabolic pathway where
the product of one enzyme becomes the substrate for the
next.
OK, lets look at how relative E & S concentrations effect what
enzymes do in a closed system (a test tube).
Enzymatic reactions take place in two steps, substrate
binding (E+S) to form an ES complex followed by catalysis,
the conversion of S to P.
ES
E+S
The E+S
While the ES
ES
E+P
binding step has a variable
speed that depends on [S] for a
given[E]
E+P
step has a speed that is
fixed by the properties of the
enzyme.
The rate of product, P, production increases as [S]
(concentration of S) increases, but only to a limit.
For each [S], the reaction rate increases to some limit; the leveling
off point for each [S] represents the equilibrium point for that [S].
You can see from this graph already that even though [S] is
increasing in even increments, the increase in rate (line slope)
is less for each successive [S] increase.
To show how [S] influences the reaction rate (speed of S
converting to P) more clearly, it helps to plot a second graph
of Velocity (rate) vs. [S]. To do this, we take data from the
first graph (rate vs. time for each [S]) shown here and plot a
second graph.
We need to select a standard time at which to compare rates represented by the vertical line drawn here. Next note the slope
of each [S] line at this point (the slope for each line expressed in
moles/min); this can be calculated from absorption readings
using the product absorption coefficient.
For each [S] the rate (tangent line slope at our selected T
value) is plotted on a new graph of velocity (slope) vs. [S]
The new graph (on the right) when complete will look
something like this.
This new graph now allows us to see other
relationships, including that an enzymatic rxn
can be saturated with S.
Increasing [S] beyond that which gives the
maximum rate or velocity, Vmax won’t
increase the rxn velocity further. How
about a simpler monkey-peanut analogy?
If you found that confusing, how about Monkeys and
Peanuts?
Lets assume that we have some monkeys and lots of peanuts,
and that we know the average time it takes for a monkey to shell
a peanut and pop it into its mouth.
This analogy has been around for awhile in various books, etc. I stole these illustrations from:
http://www.creativebiology.co.uk/#/enzyme-monkey-nuts/4546595080
So a given mix of [E] (Monkeys) and [S] (substrate) would
look like this.
Lets keep [E] (Monkeys) constant and increase [S] (peanuts). How would
this affect the time it takes to go from E+S to ES? What about the time it
takes to go from ES to E + P?
If we added more peanuts still, at some point the monkeys would have such easy
access to the peanuts that adding even more yet would not allow them to work any
faster (E+S -> ES) and we’d be at the saturation point = V max
To reiterate: At low [S] the E (monkeys), take a while to
find the few S molecules (peanuts) available in the test
tube (their cage).
As [S] (the number of peanuts) is increased, the E (monkeys),
find it easier to find S molecules (peanuts) in the test tube (their
cage) and the rate of S-to-P conversion increases.
As [S] (the number of peanuts) continues to increase (because
we are making more available) at some point, the enzyme
molecules (monkeys) simply can’t bind them any faster and
the system is saturated. Any further increase in [S] will not
increase the rate of product formation which is already at Vmax
One additional piece of info: the “km value” of an
enzyme is specific to each substrate that it can work
with.
For example our turnip peroxidase can work with a number of
materials as the non-peroxide substrate which is why the letter “R” is
used to designate it:
RH2 + H2O2
R + 2 H2 O
Some specific Kms for comparison:
A few other “Rs,” other substrates that peroxidase can also work with,
and their Km values for comparison:
Substrate
3,3’,5,5’-tetramethylbenzidine
K m*
0.045
Guaiacol
0.14
2,2’-azino-bis(3-ethylbenzthiazol-6-sulfonic acid
0.19
Phenol
1.17
O-Cresol
16.7
* Km values are mMoles of substance giving ½ Vmax under standard conditions
Km is the [S] that produces 1/2 Vmax so its reasonable that smaller
Km values indicate a tighter binding of the substrate (and the
reaction rate will be faster as a result on an equimolar basis).
Different materials (Rs) will have a
higher affinity for the enzyme = will bind
better (= lower Km), or a lower affinity for
the enzyme = looser fit (= higher Km
value). We chose guaiacol for our “R”
compound because when it reacts it
produced a colored product. The
relative km value was not an important
consideration for our purposes.
The Michaelis-Menton equation can be derived from
these values as shown; it is useful for determining how
an enzyme will behave under different conditions, but its
use will not be pursued further here.
So IGNORE THIS for the exam, it’s just an additional comment
Remember, these aspects of enzyme kinetics are
demonstrable in a closed system only - where
product cannot be removed - with an enzymatically
catalyzed reaction taking place in a test tube, such
as we did in lab for example.
In a cell, it’s a different ballgame as P is never
allowed to build up. The effects of E & S
concentration still apply however.
In a cell, enzymes are arranged into metabolic pathways
where the product of one becomes the substrate for the next.
A
Enz.1
B
C
Enz.2
D
Enz.3
E
Enz.4
F
Enz.5
Because products are removed as fast as they are made, the
materials in pathway “flow” in the forward direction. Equilibrium,
where the changes (and the accompanying energy
changes)effectively come to a standstill DOSE NOT OCCUR, if it did
it would = cellular death.
Pathways can be linear as shown (see glycolysis) or cyclic, where
F, with addition of another input, resynthesizes A and the the
pathway cycles (see the citric acid cycle).
In living cells, the product
of one enzymatic reaction
becomes the substrate for
the next.
To reiterate: At equilibrium G = 0; in living cells, reactions always
proceed in the forward direction because in a metabolic pathway
products are never allowed to accumulate, so the reverse reaction never
occurs (therefore no equilibrium point is reached). In cellular metabolism
an equilibrium state (no energy change going on) = death.
Also, a reaction, such as glucose combustion, that is carried out in
many small steps in living cells, still has the same cumulative overall
G (on an equal volume basis) as it would if you simply burned a
spoonful of it in an open flame.
Metabolic pathways can take several forms
LINEAR PATHWAY:
A
B
N
CYCLIC PATHWAY:
C
D
E
F
K
J
M
L
BRANCHING PATHWAY:
G
I
H
Intermediary metabolism
can be defined as the sum
total of all of the enzymatic
pathways that are
necessary for a cell to be
able to sustain itself.
The pathways of glycolysis,
the transition reaction, and
the TCA or Kreb’s cycle
play central “clearing
house” roles in
intermediary metabolism in
addition to being part of
aerobic respiration which
supplies ATP to the cell.
Enzymes can be inhibited (slowed down) by competitive
(substrate-like) or non-competitive (enzyme shapealtering) materials.
• A key to understanding how these processes work is
realizing that the binding between an enzyme and
it’s substrate, or an inhibitor, is seldom “tight” so it’s
an on again / off again affair that is affected greatly
by (relative) substrate and inhibitor concentrations.
• This is especially true for competitive inhibition
Figure 5.10
A competitive inhibitor has at least part
of it’s structure that is very similar in size
and shape to the size & shape of the
normal substrate. So the more Inhibitor
floating around, the harder it is for the
enzyme to find & bind to the substrate and
product production (the reaction rate)
slows down. Adding more substrate
(changing the I/S ratio in favor of S) will
speed things up again.
Non-competitive Is work by changing the overall enzyme shape, including the
shape of the active site, but because they bind at a separate allosteric site,
adding more S has no effect (won’t speed things up again).
Another view of
competitive inhibition
Another view of allosteric enzyme control. Inhibition and
activation are possible in some cases depending on which
modulators (A or I) are present.
Non-competitive inhibition is used in many cells to control the rate of an
overall pathway:
A specific example:
Isoleucine synthesis
In some cases, substrate binding (in one subunit) can
(allosterically) enhance the ease of substrate binding by
others.
Temperature and solution ion concentration can also affect
activity by affecting enzyme (and active site) 3D shape.
The shape of the enzyme and its active
site are affected by anything that alters all
of the weak interactions that determine
protein 3D shape. There is some
concentration of ions, including H+ (ie. pH)
that allows the enzyme and active site to
have their optimal shape. Under such
optimal conditions, the enzyme works the
best. Temperature also affects the rate
molecules move around and bump into
each other. So as temp. goes up, so does
reaction rate until the point where
denaturation sets in.