Aerobic Respiration: Maximizing ATP

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Aerobic Respiration Maximizing ATP
glycolysis is NOT respiration-- it is inefficient
and proceeds via direct reactions with glucose
with only low energy production respiration
oxidation driven flow of electrons through or
within a membrane, from reduced coenzymes to
an electron acceptor (oxygen) with the
generation of ATP NADH is a reduced coenzyme and
can (and does!) contribute to respiration
when it is used in fermentation (either lactate
or alcoholic) it is not taking part in
respiration fermentation has no net
oxidation/reduction-- all NAD is
regenerated oxygen is the usual ultimate
electron acceptor-- reduced oxygen is water!
ie. transfer of hydrogens along with its electrons
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Aerobic Respiration Maximizing ATP
aerobic respiration respiration in the presence
of oxygen-- complex life anaerobic respiration
respiration using other elements as electron
acceptors-- Fe3 to Fe2, sulfur S to hydrogen
sulfide H2S, a few others most aerobic ATP
production occurs in the mitochondria 36 ATP
molecules are generated overall from 1 glucose 2
of the ATP molecules are generated by glycolysis
(along with 2 NADH)
cristae are the infoldings of the inner
mitochondrial membrane creates the matrix-- area
inside both membranes and the intermembrane
space (space between the two membranes and
inside the cristae)
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Aerobic Respiration Maximizing ATP
TOM and TIM protein complexes that transport
molecules across the outer (TOM) and inner
(TIM) membranes TIM is usually found on
membranes adjacent (next to) TOM pyruvate goes
through TOM and TIM to get to the mitochondrial
matrix in the matrix pyruvate enters the TCA
(tricarboxylic acid) or Krebs cycle
4 stages of respiration (only the red one is not
in the mitochondria) 1) glycolysis--
anaerobic, serves as the entry point for the
rest 2) TCA cycle-- generation of carbon
dioxide and reduced coenzymes 3) electron
transport-- harvesting the energy of reduction
and transfer of electrons to molecular
oxygen 4) generation of ATP using the F-type
ATPase
4
Mitochondria
porins large channels in the outer mitochondrial
membrane that allow 'small' molecules (lt5000
MW) easy passage-- ATP goes right through
TOM TIM
F-type ATPases
matrix lies within the second membrane- second
membrane is not porous and ions will not flow
through directly contains no porins matrix
also contains the DNA, tRNA, etc of mitochondria
cristae are folds that point into the
mitochondria and increase the surface area
about 5x over the outside alone membranes are
required for respiration to occur
inner membrane is very dense with proteins 75
by weight! major protein of the inner
mitochondrial membrane is the F-type ATPase
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F-type ATPase
F1 complex F0 complex
matrix
side view of inner membrane
matrix inside view
F-type ATPase reversible proton pump that can
either consume or generate ATP depending upon
the direction of proton flow F-type ATPases can
also be thought of ATP synthetases composed of
6 F0 complexes on the matrix side and an F1
complex in the inner mitochondrial
membrane for prokaryotes, the cytoplasm is the
equivalent of the matrix
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TCA cycle
recall that pyruvate, in the absence of oxygen,
goes to fermentation to get rid of NADH in
the presence of oxygen, pyruvate loses a
carbon dioxide, binds coenzymeA, and reduces
a second NAD
for this reaction to occur, the pyruvate must
move to the mitochondria
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TCA cycle
acetyl-CoA is the entry route into the
tricarboxylic acid cycle tricarboxylic acid
cycle (TCA) complete oxidation of acetyl CoA to
coenzyme A and carbon dioxide also known as
the Krebs cycle after it's discoverer in the
1930's oxidative decarboxylation process of
removing a carboxyl group as carbon dioxide
and reducing a coenzyme (often NAD to
NADH) acetyl-CoA is added to oxaloacetate (the
same 4 carbon sugar from gluconeogenesis) to
form citrate, a 6 carbon tricarboxylic acid and
the beginning of the TCA cycle
CoA
8
Note the 4 reductions (!!!) and only 1 ATP
generated during the cycle
9
note that the newly added acetyl carbons are
not the ones lost during oxidative
decarboxylations they instead become part of
the oxaloacetate for the next cycle
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some cycles add another intermediate
(cis-aconitase) know where the oxidative
decarboxylations occur recognize the single
location of substrate level phosphorylation not
ice that CoA again plays a role (succinyl CoA
in converting a-ketoglutarate to succinate) 4
separate reduction reactions, 2 with oxidative
decarboxylations
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TCA cycle also leads to a number of other
important metabolic pathways
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TCA cycle
so 1 glucose to 2 molecules of acetyl- CoA gives
2 ATP and 4 NADH acetyl-CoA to CoA gives 3 NADH,
1 FADH2, and 1 GTP (ATP) overall, 1 glucose
gives 4 ATP, 10 NADH, and 2 FADH2 6 CO2 a
total of 6 carbon dioxide molecules are formed,
equal to all of the carbons in glucose we
know that carbon dioxide is the waste product--
we get rid of it nowhere is oxygen needed to
make these products! oxygen is only required
during electron transport to be the final
electron acceptor for the reduced coenzymes
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Regulating the TCA cycle
regulation of the TCA cycle is primarily
allosteric-- specific effectors (molecules)
which will activate or inhibit the enzyme by
small changes in it's protein structure 4 key
points of regulation, all using feedback
inhibition 1) pyruvate to acetyl-CoA
(generates NADH CO2) 2) isocitrate
dehydrogenase (first NADH and CO2 in the cycle
proper) 3) a ketoglutarate dehydrogenase
(second NADH and CO2) 4) malate dehydrogenase
(NADH, final step before repeating cycle) Note
the major commonalities in which enzymes are
regulated - generation of NADH (4 of 4
enzyme) - generation of CO2 (3 of 4 enzymes--
every CO2 generated!) - start (acetyl-CoA
formation) and end (malate to oxaloacetate) steps
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Regulating the TCA cycle
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Regulating the TCA cycle
negative regulators of the various enzymes
NADH (ie. when there's already a lot available,
for all 4 steps!) ATP (ie. the cell doesn't
need more energy) acetyl-CoA (for pyruvate to
acetyl-CoA, already enough product)
succinyl-CoA (feedback inhibition for making less
succinyl-CoA) negative regulators are all things
that are significant end or intermediate
products of the cycle positive regulators of the
TCA enzymes NAD (ie. need more NADH) CoA
(ie. currently low concentrations of acetyl-CoA)
ADP (ie. low energy in the cell, waiting to be
made into ATP) AMP (ie. low energy in the
cell, waiting to be made into ATP) positive
regulators are all products that are used in the
TCA cycle
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Fats feed directly into TCA Cycle
fats are the most common long term energy storage
molecules in animals store the maximum of
calories per weight, usually as triacylglycerols
each of the 3 fatty acids gets broken off the
glycerol molecule to be used uses ATP AMP
to join a fatty acid to CoA, becoming fatty
acyl-CoA one or two oxidations, depending upon
if it is saturated or unsaturated finally a 2
carbon acetyl-CoA come off and a second
molecule of CoA joins to any remaining carbon
chain of the fatty acid until there is only
acetyl-CoA left
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Fats feed directly into TCA Cycle
b oxidation of fatty acids- 2nd carbon (b) is
oxidized carbons are always removed in groups
of 2 lipids in membranes (ie. fats) are also
always found in multiples of 2 acetyl-CoA is
used for making fats and lipids as well as
breaking them down-- it is a central molecule
in all of cellular metabolism
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Proteins feed into the TCA Cycle
proteins are much less likely to be used for
energy than fats and sugars they take too much
energy to build to be wasted that
way! proteolysis breakdown of proteins by
various enzymes (proteases) the big difference
between proteins and fats/sugars is the amine
group
O
O
O
O
O
O
O
O








R-CH-C-O- -O-C-C-CH2-CH2-C-O-
R-C-C-O- -O-C-CH-CH2-CH2-C-O-


NH3
NH3
a-ketoglutarate (TCA cycle)
keto acid easy to convert to intermediate
glutamate (amino acid)
general amino acid
several amino acids can convert directly into TCA
cycle intermediates glutamate to a-ketoglutarate,
alanine to pyruvate, aspartate to oxaloacetate
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Proteins feed into the TCA Cycle
glutamate and a few others can lose amines by
oxidative deamination
O
O


-O-C-CH-CH2-CH2-C-O- NAD H2O NH4
NADH H a-ketoglutarate

NH3
note that transaminations are reversible-- TCA
intermediates can be used to build amino acids
as well as break them down for energy TCA cycle
serves at the center of a complex series of
reactions that are used to make up amino acid
monomers, lipids, etc that the cell needs the
tricarboxylic acid cycle is at the center of a
very large series of these metabolic reactions
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Electron transport chain Oxygen gets its
electrons
from one molecule of glucose, we've generated a
LOT of reduced coenzymes but very very little
actual ATP the body has relatively few coenzyme
molecules-- most are derivatives of the B
vitamins (niacin - nicotinamide, flavin (FADH)-
riboflavin) oxygen has yet to make an
appearance-- we have to take the electrons
from the coenzymes and move them to molecular
oxygen while coupling that energy for
metabolism DG values for the reduction of NADH
and FADH2 are enormous NADH -52.4 kcal/mol,
FADH2 -45.9 kcal/mol to couple as much of
that energy as possible, it must go through
multiple steps-- direct conversion to ATP
(-7.3 kcal/mol) would be inefficient
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Electron transport chain Oxygen gets its
electrons
electron transport system (ETS) series of
coenzymes coupled to proteins in membranes
which move electrons to molecular oxygen while
using the energy to create a proton gradient
across the inner mitochondrial membrane 5
different carriers in the electron transport
chain flavoproteins iron-sulfur proteins
cytochromes copper containing cytochromes
coenzyme Q respiratory complexes large
groupings of various members of the electron
transport chain most electron transport system
molecules also absorb light of particular
wavelengths, giving them particular colors
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Electron Transport Chain Flavoproteins
always has a prosthetic group to carry the
electrons-- either flavin adenine dinucleotide
or flavin mononucleotide transfers both
electrons and protons as they are oxidized or
reduced work as catalysts like other enzymes
do not change after reacting includes NADH
dehydrogenase, the enzyme that regenerates NAD
23
Electron Transport Chain Iron-sulfur Proteins
have an ironsulfur cluster that is complexed
to the protein by cysteines iron serves as the
electron carrier cycling between Fe3
(oxidized) and Fe2 (reduced) most numerous
type of protein in the mitochondrial electron
transport chain with 12 members unlike NADH,
iron-sulfur proteins move one electron iron-sulfu
r proteins also do not pick up or release protons
when moving their electrons around
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Electron Transport Chain Cytochromes
cytochromes also contain iron but with a heme
prosthetic group-- ie. like hemoglobin 5
cytochromes in the electron transport chain,
some with slightly modified heme groups iron
serves as the electron acceptor cytochrome C
serves to transfer electrons between complexes
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Electron Transport Chain Copper Cytochromes
iron heme
copper- heme
copper cytochromes contain 2 copper ions instead
of iron can also exist as an iron-copper center
(ie. one iron, 1 copper) in heme like iron,
copper can be oxidized (Cu2) or reduced
(Cu) iron copper center is critical for holding
onto the O2 molecule until it acquires 4
electrons (and protons! H) to form 2 water (H2O)
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Electron Transport Chain Coenzyme Q
also known as ubiquinone can move 1 or 2
electrons (and protons) as semiquinone (1 e-)
or dihydroquinone (2e-) ketone oxygens are the
atoms that pick up the hydrogens during
reduction exists by itself in the inner
mitochondrial membrane-- hydrophobic tail
allows it to diffuse freely within the
membrane carries electrons between other
carriers and other respiratory complexes most
abundant electron carrier in the membrane (hence
its name) coenzyme Q is vital for pumping
hydrogens across the membrane-- acepts
hydrogens in the matrix and releases them in the
intermembrane space, resulting in them moving
across the membrane
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Electron Transport Chain Sequence of Transfers
reduction potential (redox potential) measure of
the relative reducing power of a particular
redox pair-- how strongly does the oxidized
state want to hold onto its electrons redox
pair given ions or chemicals which switch
between oxidized and reduced states always
written as oxidized/ reduced forms, as in
NAD/NADH positive redox potential really
wants to keep it's electrons-- ie.
oxygen negative redox potential doesn't mind
giving up electrons-- ie. NAD more negative
redox potentials will give up their electrons to
more positive ones-- this determines the
direction of the electron flow
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Electron Transport Chain Redox Potentials
standard reduction potential Eo', reduction
potential for a redox pair under standard
conditions (25 oC, 1M concentrations, pH 7.0)
table of standard reduction potentials allow
you to easily determine if a compound will
oxidize or reduce another compound on the
list the reduced form will spontaneously
reduce compounds below it on the table-- ie.
NADH will reduce oxygen to water, cytochrome b
will reduce cytochrome c, etc.
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Electron Transport Chain Redox Potentials
you can calculate the actual reduction potential
using E' Eo' RT/nF ln oxidized/reduced
where F faraday's constant n electrons
transferred, and R gas constant note that E'
is still at pH7.0-- that's what the '
means! DEo' Eo'acceptor -Eo'donor (note
spontaneous reaction is !!!) you can also
calculate DGo' from DEo' DGo' -nFDEo' so you
can calculate the total change in free energy for
moving any given distance along the standard
reduction potential ie. NADH to oxygen
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Electron Transport Chain Redox Potentials
What is DGo' for NADH reducing O2? DGo' -nF
DEo' DGo' -nF (Eo'oxygen-Eo'NADH) DGo'
-223062 (0.82- (-0.32)) DGo' -52.4 kcal/mol
in the electron transport chain going from NADH
to oxygen, each link in the chain must be at a
lower reduction potential than the one above
it This will make DG negative and allow the
electrons to move spontaneously to reach oxygen
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Electron Transport Chain Redox Potentials
respiratory complex one of 4 different complexes
containing the enzymes of the electron
transport chain complex I starts with NADH and
moves electrons to coenzyme Q complex II starts
with FADH2 and moves electrons to coenzyme
Q complexes I and II start at different places
(NADH is higher than FADH2 on the table) and
therefore require different pathways to get the
max energy out of them after reaching coenzyme
Q, the NADH and FADH2 pathways intersect complex
III starts with coenzyme Q and moves electrons
to cytochrome c complex IV starts with
cytochrome c and moves electrons to O2 (end)
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Electron Transport Chain Redox Potentials
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Electron Transport Chain Redox Potentials
each complex has multiple proteins with various
prosthetic groups and different electron
potentials each complex also pumps various
numbers of hydrogens across the membrane I
pumps 4, CoQ pumps 2, III pumps 2, and IV pumps
2 complex II doesn't move any protons across
the membrane thus each NADH moves 10 H ions and
each FADH2 moves 6 H across the membrane to
make ATP by the F-type ATPase the final enzyme
in complex IV is cytochrome C oxidase that is
the terminal oxidase-- transfers electrons to
molecular oxygen azide and cyanide, two nasty
poisons, target this enzyme and inhibit it,
eventually stopping the entire electron transport
chain because it must go through cytochrome c
oxidase
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pumping protons back the F-type ATPase
F1 complex F0 complex
matrix
side view of inner membrane
matrix inside view
think of the F-type ATPase as a turbine-- the F0
subunits 'turn' using the energy of H ions
flowing across the membrane generating ATP H
flows through the F1 complex, into F0, and back
into the matrix 3 H moving across the membrane
enough to generate 1 ATP from ADP
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pumping protons back the F-type ATPase
a1
a3
a2
a1
b3
b2
b1
b3
b1
b3
b2
b1
a3
a2
a1
a3
a2
a1
a3
a2
b2
b1
b3
b2
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
P ADP ATP
as the F0 subunits rotate, they must undergo
allosteric structural changes those changes
are what drive the addition of a phospate to ADP
making ATP-- chemiosmotic model of ATP
synthesis