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Making energy!
ATP
The point
is to make
ATP!
Enduring Understandings
 II.
A. Growth, reproduction and maintenance of the
organization of living systems require free energy and
matter.
1.
2.
 IV.
All living systems require a constant input of free energy.
Organisms capture and store free energy for use in biological
processes.
A. Interactions within biological systems lead to
complex properties.
2.
The structure and function of subcellular components, and their
interactions, provide essential cellular processes.
Discussion
 What is the fundamental coupled reaction
that makes up cellular respiration?
The energy needs of life
 Organisms are endergonic systems

Humans require ~2000 kilocalories
per day

What do we need energy for?
 synthesis
 building biomolecules
 reproduction
 movement
 active transport
 temperature regulation
Where do we get the energy from?
 Work of life is done by energy coupling

use exergonic (catabolic) reactions to fuel
endergonic (anabolic) reactions
digestion
+
synthesis
+
+
energy
+
energy
Recall ATP…
ADP
ATP
O– O– O –
–O P –O
O– P –O
O– P O–
O O O
O–
–O P O – +
O
7.3
energy
 ATP  ADP

releases energy
 ∆G = -7.3 kcal/mole
 Fuel other reactions
 Phosphorylation

released Pi can transfer to other molecules
 destabilizing the other molecules

enzyme that phosphorylates = “kinase”
An example of Phosphorylation:
dehydration synthesis
 Building polymers from monomers


H H
C C
OHHO
need to destabilize the monomers, H2O
doesn’t just come off on its own
phosphorylate!
H
C
OH
H
C
OH
H
C
+
H
C
HO
synthesis
+4.2 kcal/mol
“kinase”
+ ATP
+
P
H
C
HO
enzyme
-7.3 kcal/mol
-3.1 kcal/mol
enzyme
H H
C C
O
H
C
+
+
H2O
ADP
P
H H
C C
O
+
Pi
Another example of Phosphorylation…
 The first steps of cellular respiration
starting the breakdown of glucose requires some
ATP investment
 “Substrate-level phosphorylation”

glucose
C-C-C-C-C-C
hexokinase
phosphofructokinase
P
2 ATP
C
C
2 ADP
fructose-1,6bP
P-C-C-C-C-C-C-P
DHAP
P-C-C-C
G3P
C-C-C-P
H
C
P
activation
energy
H+
H+
Our end goal…
H+
 ATP Synthase, enzyme
H+
H+
H+
H+
H+
channel in
mitochondrial
membrane


rotor
permeable to H+
H+ flow down
concentration gradient
rod
catalytic
head
 flow like water over
water wheel
 flowing H+ cause
change in shape of
ATP synthase enzyme
 powers bonding of
Pi to ADP:
ADP + Pi  ATP
ADP + P
ATP
H+
Cellular Respiration
Harvesting Chemical Energy
ATP
Harvesting stored energy
 Energy is stored in organic molecules
carbohydrates, fats, proteins
Heterotrophs eat these organic molecules  food
 Catabolize/digest organic molecules to get…


 raw materials for synthesis
 fuels for energy
 controlled release of energy
 “burning” fuels in a series of
step-by-step enzyme-controlled reactions
Overview of cellular respiration
 4 metabolic stages

Anaerobic respiration
1. Glycolysis
 respiration without O2
 in cytosol

Aerobic respiration
 respiration using O2
 in mitochondria
2. Pyruvate oxidation
3. Krebs cycle
4. Electron transport chain
->ATP Synthase
C6H12O6 +
6O2
 ATP + 6H2O + 6CO2 (+ heat)
Cellular Respiration
Stage 1:
Glycolysis
Glycolysis
 Breaking down glucose

“glyco – lysis” (splitting sugar)
glucose      pyruvate
2x 3C
6C

ancient pathway which harvests energy
 where energy transfer first evolved
 transfer energy from organic molecules to ATP
 still is starting point for ALL cellular respiration

but it’s inefficient
 generate only 2 ATP for every 1 glucose

Anaerobic, occurs in cytosol
That’s
not enough
ATP for me!
Discussion
 Why does it make evolutionary sense that
the earliest of the energy-releasing
processes is glycolysis, which takes place
in the cytosol?
Evolutionary perspective
glucose
C-C-C-C-C-C
Overview
10 reactions

convert
glucose (6C) to
2 pyruvate (3C)
 produces:
+4 ATP & +2
NADH
 consumes:
-2 ATP

enzyme
2 ATP
enzyme
2 ADP
fructose-1,6bP
P-C-C-C-C-C-C-P
enzyme
enzyme
enzyme
DHAP
P-C-C-C
net yield:
2 pyruvate, 2 ATP
& 2 NADH
DHAP = dihydroxyacetone phosphate
G3P = glyceraldehyde-3-phosphate
G3P
C-C-C-P
2H
2Pi enzyme
2 NAD+
2
enzyme
2Pi
4 ADP
enzyme
pyruvate
C-C-C
4 ATP
Glycolysis summary
endergonic
invest some ATP
ENERGY INVESTMENT
-2 ATP
ENERGY PAYOFF
G3P
C-C-C-P
4 ATP
exergonic
harvest a little
ATP & a little NADH
like $$
in the
bank
NET YIELD
net yield
2 ATP
2 NADH
Is that all there is?
 Not a lot of energy…

for 1 billon years+ this is how life on
Earth survived
 no O2 = slow growth, slow reproduction
 only harvest 3.5% of energy stored in glucose
 more carbons to strip off = more energy to harvest
O2
O2
O2
O2
O2
glucose     pyruvate
2x 3C
6C
But can’t stop there!
G3P
DHAP
NAD+
raw materials  products
Pi
+
NADH
NAD
NADH
Pi
1,3-BPG
NAD+
Pi
+
NADH
NAD
1,3-BPG
NADH
7
ADP
Glycolysis
6
Pi
ADP
ATP
ATP
3-Phosphoglycerate
(3PG)
3-Phosphoglycerate
(3PG)
2-Phosphoglycerate
(2PG)
2-Phosphoglycerate
(2PG)
glucose + 2ADP + 2Pi + 2 NAD+  2 pyruvate + 2ATP
+ 2NADH
8
 Going to run out of NAD+


9
H2O
without regenerating NAD+,
energy production would stop! Phosphoenolpyruvate
(PEP)
another molecule must accept e-ADP
10
from NADH
ATP
 so NAD+ is freed up for another round
Pyruvate
H2O
Phosphoenolpyruvate
(PEP)
ADP
ATP
Pyruvate
How is NADH recycled to NAD+?
Another molecule
must accept H
from NADH
H2O
O2
recycle
NADH
with oxygen
without oxygen
aerobic respiration
anaerobic respiration
“fermentation”
pyruvate
NAD+
NADH
acetyl-CoA
CO2
NADH
NAD+
lactate
acetaldehyde
NADH
NAD+
lactic acid
fermentation
which path you
use depends on
who you are…
Krebs
cycle
ethanol
alcohol
fermentation
Pyruvate is a branching point
Pyruvate
O2
O2
fermentation
anaerobic
respiration
mitochondria
Krebs cycle
aerobic respiration
Fermentation (anaerobic)
 Yeast, fungi
pyruvate  ethanol + CO2
3C
NADH
2C
NAD+
 beer, wine, bread
1C
back to glycolysis
 Animals, some bacteria
pyruvate  lactic acid
3C
NADH
3C
NAD+back to glycolysis
 cheese, anaerobic exercise (no O2)
Alcohol Fermentation
pyruvate  ethanol + CO2
3C
NADH
2C
1C
NAD+ back to glycolysis
 Dead end process
 at ~12% ethanol,
kills yeast
 can’t reverse the
reaction
Bacteria
Fungi
recycle
NADH
Lactic Acid Fermentation
pyruvate  lactic acid

3C
NADH
O2
3C
NAD+ back to glycolysis
 Reversible process
 once O2 is available,
lactate is converted
back to pyruvate by
the liver
 Why would this be
reversible but not
alcoholic ferm.?
(Hint: C)
animals
bacteria
recycle
NADH
Discussion
 Knowing what you do about glucose
catabolism so far, how can we use bacteria
and yeast to…
Make bread rise?
 Make alcoholic drinks?


If you want to make bread or an alcoholic
drink, what should the bacteria or yeast
environment contain? What should it not
contain?
Cellular Respiration
Stage 2 & 3:
Oxidation of Pyruvate
Krebs Cycle
Mitochondria — Structure
 Double membrane energy harvesting organelle


smooth outer membrane
highly folded inner membrane
 cristae

intermembrane space
 fluid-filled space between membranes

matrix
 inner fluid-filled space


prokaryotic DNA (mDNA), ribosomes
enzymes
intermembrane
 free in matrix & membrane-bound space
What cells would have
a lot of mitochondria?
outer
membrane
inner
membrane
cristae
matrix
mitochondrial
DNA
Mitochondria – Function
Oooooh!
Form fits
function!
Membrane-bound proteins: Enzymes & permeases
(membrane transport proteins)
Advantage of highly folded inner membrane?
More surface area for membrane-bound enzymes & permeases
Discussion
 Thinking of the human body, which kinds of
cells would you expect would have more
mitochondria? Which would you expect
would have less? (If you’ve learned this in
anatomy, be nice, give your partner a
chance to try their hand at it first :P)
Glycolysis is only the start
 Glycolysis
glucose      pyruvate
6C
2x 3C
 Pyruvate has more energy to yield
3 more C to strip off (to oxidize)
 if O2 is available, pyruvate enters
mitochondria
 enzymes of Krebs cycle complete the full
oxidation of sugar to CO2

pyruvate       CO2
3C
1C
Cellular respiration
Oxidation of pyruvate
 Pyruvate enters mitochondrial matrix
[
2x pyruvate    acetyl CoA + CO2
3C
2C
1C
NAD

Pyruvate is oxidized
 releases 2 CO2 (count the carbons!)
 reduces 2 NAD  2 NADH (moves e-)
 produces 2 (two-carbon) acetyl CoA
 Acetyl CoA enters Krebs cycle
]
Pyruvate oxidized to Acetyl CoA
reduction
NAD+
Pyruvate
C-C-C
[
Coenzyme A
CO2
Acetyl CoA
C-C
oxidation
2 x Yield = 2C sugar + NADH + CO2
]
Krebs cycle
1937 | 1953
 aka Citric Acid Cycle
in mitochondrial matrix
 8 step pathway

 each catalyzed by specific enzyme
Hans Krebs
1900-1981
 step-wise catabolism of 6C citrate molecule
 Evolved later than glycolysis

Evolutionarily…
 bacteria 3.5 billion years ago (glycolysis)
 free O2 2.7 billion years ago (photosynthesis)
 eukaryotes 1.5 billion years ago (aerobic
respiration = organelles  mitochondria)
Count the carbons!
pyruvate
3C
2C
6C
4C
This happens
twice for each
glucose
molecule,
because
glycolysis
produced two
pyruvates
4C
acetyl CoA
citrate
oxidation
of sugars
CO2
x2
4C
4C
6C
5C
4C
CO2
Count the electron carriers!
pyruvate
3C
2C
6C
4C
NADH
4C
4C
acetyl CoA
citrate
reduction
of electron
carriers
x2
FADH2
4C ATP
CO2
NADH
6C
CO2
NADH
5C
4C
CO2
NADH
What happened?
So we fully
oxidized
glucose
C6H12O6

CO2
& ended up
with 4 ATP!
What’s the
point? :/
http://highered.mcgraw-hill.com/sites/0072507470/student_view0/chapter25/animation__how_the_krebs_cycle_works__quiz_1_.html
Electron Carriers = Hydrogen Carriers
H+
 Krebs cycle
produces large
quantities of
electron carriers
NADH
 FADH2
 go to Electron
Transport Chain!

H+
H+
H+
+
H+ H H+
H+
ADP
+ Pi
ATP
H+
Discussion
 Krebs Cycle and Glycolysis have given us


energy carriers NADH, FADH2…
…which will go to the electron transport
chain…
…where ATP synthase is located…
 PREDICT… how will we be able to use
NADH and FADH2 to make ATP??
Energy accounting of Krebs cycle
4 NAD + 1 FAD
4 NADH + 1 FADH2
2x pyruvate          CO2
3C
3x 1C
1 ADP
1 ATP
ATP
Net gain = 2 ATP
= 8 NADH + 2 FADH2
Cellular Respiration
Stage 4:
Electron Transport Chain
Cellular respiration
ATP accounting so far…
 Glycolysis  2 ATP
 Kreb’s cycle  2 ATP
 Life takes a lot of energy to run, need to
extract more energy than 4 ATP!
There’s got to be a better way!
I need a lot
more ATP!
A working muscle recycles over
10 million ATPs per second
There is a better way!
 Electron Transport Chain

series of proteins built into
inner mitochondrial membrane
 along cristae
 transport proteins & enzymes
transport of electrons down ETC linked to
pumping of H+ to create H+ gradient
 yields ~36 ATP from 1 glucose!
 only in presence of O2 (aerobic respiration)

That
sounds more
like it!
O2
Remember: Mitochondria
 Double membrane


outer membrane
inner membrane
 highly folded cristae
 enzymes & transport proteins
 Matrix space within the inner
membrane

intermembrane space
 fluid-filled space between
membranes
Electron Transport Chain
Intermembrane space
Outer mitochondrial
membrane
Inner
mitochondrial
membrane
C
Q
NADH
dehydrogenase
cytochrome
bc complex
Mitochondrial matrix
cytochrome c
oxidase complex
Remember the Electron Carriers?
Glycolysis
glucose
Krebs cycle
G3P
2 NADH
Time to
break open
the piggybank!
8 NADH
2 FADH2
Electron Transport Chain
Building proton gradient!
NADH  NAD+ + H
e
p
intermembrane
space
H+
H+
H  e- + H+
H+
C
e–
Q
e–
NADH H
FADH2
NAD+
NADH
dehydrogenase
inner
mitochondrial
membrane
e–
H
FAD
2H+ +
cytochrome
bc complex
1
2
O2
H2O
cytochrome c
oxidase complex
mitochondrial
matrix
What powers the proton (H+) pumps?…
Stripping H from Electron Carriers
 Electron carriers pass electrons to ETC

electrons stripped from H atoms
 electrons passed from one electron carrier to next in
mitochondrial membrane (ETC)
 flowing electrons = energy to do work

transport proteins in membrane pump H+ (protons)
across inner membrane to intermembrane space
H+
+
H
H+
TA-DA!!
Moving electrons
do the work!
+
H
H+
+
H
H+
H+
+
H+ H+ H
+
H+ H H+
C
e–
NADH
Q
e–
FADH2
FAD
NAD+
NADH
dehydrogenase
e–
2H+
cytochrome
bc complex
+
1
2
O2
H2O
cytochrome c
oxidase complex
ADP
+ Pi
ATP
H+
But what “pulls” the
electrons down the ETC?
H 2O
O2
electrons
flow downhill
to O2
oxidative phosphorylation
Electrons flow downhill
 Electrons move in steps from
carrier to carrier downhill to oxygen
each carrier more electronegative
 controlled oxidation
 controlled release of energy

Taking it beyond…
 What is the final
H+
H+
H+
C
Q
e–
FADH2
FAD
electron acceptor in
e
NADH
2H +
NAD
Electron Transport
O2
Chain?
 So what happens if O2 unavailable?
 ETC backs up
e–
–
+
+
NADH
dehydrogenase
cytochrome
bc complex
1
2
O2
H2O
cytochrome c
oxidase complex
nothing to pull electrons down chain
 NADH & FADH2 can’t unload H

 ATP production ceases
 cells run out of energy
 and you die!
Pyruvate from
cytoplasm
Inner
+
mitochondrial H
membrane
H+
Intermembrane
space
Electron
transport
C system
Q
NADH
Acetyl-CoA
1. Electrons are harvested
and carried to the
transport system.
NADH
Krebs
cycle
e-
e-
FADH2
e-
2. Electrons
provide energy
to pump
protons across
the membrane.
e-
H2O
3. Oxygen joins
with protons to
form water.
1 O
2 +2
2H+
O2
H+
CO2
ATP
Mitochondrial
matrix
H+
ATP
ATP
4. Protons diffuse back in
down their concentration
gradient, driving the
synthesis of ATP.
H+
ATP
synthase
http://www.qcc.cuny.edu/biologicalsciences/Faculty/UGolebiewska/respiration.html
://www.youtube.com/watch?v=FFBr3ANCkb4
Cellular respiration
2 ATP
+
2 ATP
+
~36 ATP
Discussion
C6H12O6 + 6O2







 6CO2 + 6H2O + ~40 ATP
Where did the glucose come from?
Where did the O2 come from?
Where did the CO2 come from?
Where did the CO2 go?
Where did the H2O come from?
Where did the ATP come from?
What else is produced that is not listed
in this equation?
 Why do we breathe?
 http://www.youtube.com/watch?v=FFBr3AN
Ckb4
Comparison of Chemiosmosis in
Chloroplasts and Mitochondria
ETC of Photosynthesis
Chloroplasts transform light energy
into chemical energy of ATP

generates O2
use electron carrier NADPH
ETC of Respiration
Mitochondria transfer chemical energy from food molecules
into chemical energy of ATP

use electron carrier NADH
generates H2O
Cellular Respiration
Other Metabolites &
Control of Respiration
Cellular respiration
Other metabolites
 Cellular respiration is sequential. We can
enter at multiple points along the pathway,
if we can produce the molecule that usually
belongs there some other way.

We don’t HAVE to start with pure glucose!
Beyond glucose: Other carbohydrates
 Glycolysis accepts a wide range of
carbohydrates fuels
polysaccharides    glucose
hydrolysis
 ex. starch, glycogen
other 6C sugars    glucose
modified
 ex. galactose, fructose
Beyond glucose: Fats
fats      glycerol + fatty acids
hydrolysis
glycerol (3C)   G3P   glycolysis
fatty acids  2C acetyl  acetyl  Krebs
groups
coA
cycle
3C glycerol
enters
glycolysis
as G3P
2C fatty acids
enter
Krebs cycle
as acetyl CoA
Carbohydrates vs. Fats
 Fat generates 2x ATP vs. carbohydrate

more C in gram of fat
 more energy releasing bonds

more O in gram of carbohydrate
That’s why
it takes so much
work to lose a
pound a fat!
 already partly oxidized
 less energy to release
carbohydrate
fat
Discussion
 Why would an organism ever convert acetyl
CoA to fat instead of putting it through the
Krebs Cycle and ETC? Krebs + ETC = ATP!
ATP powers everything! What gives?
Only 2C, clearly not a lot of
energy this way, cells try to
avoid it
Beyond glucose: Proteins
proteins      amino acids
hydrolysis
waste
H O
H
| ||
N —C— C—OH
|
H
R
amino group =
waste product
excreted as
ammonia, urea,
or uric acid
glycolysis
Krebs cycle
2C sugar =
carbon skeleton =
enters glycolysis
or Krebs cycle at
different stages
Metabolism
 Digestion

digestion of
carbohydrates, fats &
proteins
 all catabolized through
same pathways
 enter at different points

cell extracts energy
from every source
Cells are
versatile
&
CO2
selfish!
Metabolism
 Synthesis


enough energy?
build stuff!
cell uses points in
glycolysis & Krebs cycle
as links to pathways for
synthesis
 run pathways “backwards”
 have extra fuel, build fat!
pyruvate
  glucose
Krebs cycle
intermediaries
acetyl CoA

amino
acids
  fatty acids
Cells are
versatile &
thrifty!
Metabolic Rate
Metabolic Rate
 Metabolic rate is the amount of energy
required by an organism.

Usually measured in units of energy per
body mass per time (such as J/g/hr), or
simply in energy per time (such as mm
O2/day).
Metabolic Rate
 Think of examples of animals of all sizes,
and then hypothesize: who do you think
probably has a higher metabolic rate
(kcal/g/day), smaller organisms, larger
ones, or no relationship? (Hint: it takes
energy to maintain body temperature)
Metabolic Rate
 Living things
must expend
energy to
maintain the right
body temperature
for these enzymes
to function
Metabolic Rate
Metabolic Rate
 Relationship exists between
thermoregulation and
metabolic rate


Endotherms (mammals, birds) =
Temperature regulation mostly
thanks to internal processes: heat
lost during routine metabolic
processes
Ectotherms (all other animals) =
Body temperature regulation
mostly thanks to external
environment: very little heat
generated by normal metabolism
Discussion
 Predict: Sketch a graph with two lines: one
for endotherms, one for ecotherms.
X-Axis: Ambient (environmental)
temperature
 Y-Axis: Rate of oxygen consumption

Generating Heat
 Oxidative phosphorylation can be
decoupled from the ETC
Decoupling protein can redirect protons
back across the membrane
 No ATP formed
 Energy dissipated as heat instead
