Mitochondria, Peroxisomes and Chloroplasts

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Mitochondria, Peroxisomes andChloroplasts

• Dr. Bill Diehl-Jones
• 22.228

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Agenda

• Metabolism
• A general review
• Catabolism and Anabolism
• Oxidation and Reduction
• The Mitochondrion
• Peroxisomes
• Chloroplasts

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Metabolism
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Overview Metabolism - sum total of all chemical
changes that occur in cells. - each reaction
is catalyzed by a specific enzyme. - compounds
formed in each step along the pathway are
metabolites. - pathway leads to an
endproduct. - endproduct has a particular role
in the cell. - two broad types of metabolic
pathways catabolic and anabolic
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1. Catabolic pathways Greek prefix -kata
down - breaking of chemical bonds in large,
complex molecules to form small simple molecules.
- these molecules can be used to synthesize
other molecules. - catabolic reactions are
exergonic. - provide chemical energy for cell
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2. Anabolic pathways - Greek prefix -ana
up - synthesis of large molecules by chemically
bonding together small molecules. - anabolic
reactions are endergonic. - require
energy Catabolic and anabolic pathways are
interconnected. - catabolic pathways provide
energy and small molecules for anabolic pathways.
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Glycolysis and Gluconeogenesis - catabolic
(glycolysis) and anabolic (gluconeogenesis)
pathways for the metabolism of glucose 1.
Glycolysis a. a universal catabolic
pathway. b. breakdown of glucose. c. ten-step
reaction sequence (glucose to pyruvate). d.
occurs in presence or absence of O2. e. occurs
in cytosol.
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Glycolytic Pathway Chart
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2 Glycolytic products for 1 molecule of
glucose a. uses 2 ATP and produces 4 ATP - ATP
formed by substrate-level phosphorylation b.
yields 2 pyruvates - pyruvate stands at junction
of anaerobic vs aerobic metabolism c. yields
2 NADH i. fermentation ii. electron transport
chain in mitochondria iii. make NADPH
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3. Reducing Power - a cells reservoir of
NADPH. - formation of complex biological
molecules requires reduction of precursors. -
accomplished by the energy-releasing transfer of
electrons from NADPH (the electrons themselves
are not high E). - NADPH is coenzyme for
enzymes having reductive role in anabolic
pathways. e.g. C3 cycle in photosynthesis e.g.
fat - NADPH is interconvertible with NADH.
(Under special circumstances). - NAD is
coenzyme for dehydrogenases in catabolic
pathways. e.g. glyceraldehyde-3-phosphate
dehydrogenase in glycolysis
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4. Equilibrium versus- steady state metabolism -
many reactions in a metabolic pathway may be near
equilibrium. - however, several reactions in a
pathway are poised far from equilibrium. - these
are essentially irreversible and keep pathway
going in a single direction. - cellular
metabolism can maintain itself at irreversible
nonequilibrium conditions because the cell is an
open system. - an open system because materials
are continually flowing in and out of the
cell. - cellular metabolism is said to exist in
a steady state. - in a steady state, the
concentrations of reactants and products remain
essentially constant, even though individual
reactions are not necessarily at equilibrium.
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• 5. Driving force for glycolysis
• when concentration of metabolites in the cell are
  measured, three reactions are far from
  equilibrium
• 1. hexokinase
• 2. phosphofructokinase
• 3. pyruvate kinase
• - these 3 are essentially irreversible and keep
  pathway going in a single direction.
• - All subject to feedback inhibition by ATP or
  other factors

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hexokinase
phosphofructokinase
pyruvate kinase
Free-energy changes of glycolytic reactions
under cellular (non-standard) conditions
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6. Separating Catabolic and Anabolic Pathways
1. e.g. glycolysis and gluconeogenesis
glycolysis (catabolic) - breakdown of glucose
gluconeogenesis (anabolic) - formation of
glucose 2. thermodynamic problem - cannot
proceed simply by reversal of reactions. glycoly
tic pathway contains 3 thermodynamically
irreversible reactions. 3. regulatory problem -
two pathways could not be controlled
independently of one another.
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7. Separating glycolysis and gluconeogenesis 1.
Solved by using different enzymes to catalyze 3
key reactions in 2 opposing pathways. e.g.
step between fructose-6-phosphate and fructose
1,6-bisphosphate 2. Other reactions are
identical, although they run in opposite
directions.
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8. Phosphofructokinase in glycolysis Fructose
6-phosphate ATP ? ADP Fructose 1,
6-bisphosphate - This is an allosteric enzyme.
positive modulators ADP, AMP negative
modulator ATP - When cell has ample ATP,
the enzyme is inhibited. - If ATP is being used
up, the enzyme is stimulated.
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Feedback Inhibition
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9. Fructose 1,6-bisphosphatase in
gluconeogenesis Fructose 1,6-bisphosphate H2O
? Fructose 6-phosphate Pi - This is a
covalently modified enzyme. - When glucose
levels are high, enzyme is inhibited. - This is
done by phosphorylation of an amino acid
residue. Regulating ATP levels - Generally
ATP levels do not fluctuate - ATP must be
maintained high relative to that of ADP and
AMP. - In this way, -?G of ATP hydrolysis
remains large enough to drive endergonic
reactions.
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Hydrolysis of ATP
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Aerobic Respiration   1. Aerobic oxidation of
pyruvate (  1. When O2 is present, pyruvate
enters mitochondria.   2. completely oxidized to
CO2.   a. pyruvate dehydrogenase   b. TCA or
Krebs cycle   3. get up to 36 ATP/glucose
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a. Oxidation of pyruvic acid to acetyl CoA   1.
Catalyzed by pruvate dehydrogenase complex 2.
One CO2 is evolved.   3. NADH is formed
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b. TCA or Krebs cycle   1. 2-carbon acetyl
group condenses with 4-carbon oxaloacetate to
form 6-carbon citrate.   2. one turn of cycle
evolves 2 CO2.   3. this completes oxidation of
pyruvate.   4. free energy is conserved in   a.
3 NADH and 1 FADH2. - will be used in
electron-transport chain to form ATP.   b. 1 GTP
- energetically equivalent to ATP
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c. TCA or Krebs cycle as central metabolic
pathway of cell   1. metabolites of other
catabolic pathways enter TCA cycle e.g.
breakdown of proteins   2. TCA cycle metabolites
can be used to synthesize larger molecules
(anabolism). e.g. amino acids for protein
synthesis   3. amphibolic pathway - used in both
catabolism and anabolism.
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d. Structure of the Mitochondrion   1. outer
membrane i. porins - integral proteins that form
large, nonselective membrane channels.   2.
intermembrane space   3. inner membrane i.
electron transport chain ii. ATP synthase   4.
matrix i. TCA cycle ii. DNA (genes for 13
polypeptides) iii. ribosomes
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Mitochondria

• Visible by light-microscopy,
• 1500 per liver cell, 15 - 20 of the cell volume
• Production of ATP
• Structure
• Outer membrane, inner membrane (folded into
  cristae), inter-membrane space, matrix.

http//tidepool.st.usm.edu/pix/mitochondrion.gif
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Inter-membrane space
O.M.M.
Cristae
I.M.M.
DNA strand
matrix
ribosomes
ATP synthase particles
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2. Electron transport chain   - sequential
transfer of electrons from one electron carrier
to another. - all carriers are bound in inner
mitochondrial membrane.   - four types of
electron carriers
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• a. Components of electron transport chain
•  
• Flavoproteins
• - prosthetic groups are derived from riboflavin.
  (FAD
• or FMN) e.g. NADH dehydrogenase
•  
• 2. cytochromes
•   - proteins that contain heme groups.
• - iron of heme undergoes reversible transition
• between Fe3 and Fe2 .
•  
• 3. ubiquinone (or coenzyme Q)
•   - lipid-soluble molecule
•  
• 4. iron-sulfur proteins
•  - iron is closely linked to inorganic sulfur.

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b. Function of Electron Transport Chain   1.
Free energy released from oxidation/reductions of
electron transport chain - moves H from matrix
to intermembrane space.   2. This sets up a H
gradient.   3. Ionic gradient across a membrane
represents a form of energy.   4. Energy is used
to form ATP. This is oxidative phosphorylation.
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Electron Transport Chaincreates hydrogen ion
gradient, consumes oxygen
inter-membrane space
H
H
H
IV
III
I
II
2H 1/2 O2
H2O
2 e-
2 e-
O2 has a high electron affinity
NADH NAD
FADH2 FAD 2H
low electron affinity
II. succinate dehydrogenase
I. NADH Dehydrogenase
III. cytochrome bc1
IV. cytochrome oxidase
matrix
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Electron Transport Portion of Energy Metabolism
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Electron Carriers of Respiratory Chain
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Key Points

• Protons are translocated across the membrane,
  from the matrix to the intermembrane space
• Electrons are transported along the membrane,
  through a series of protein carriers
• Oxygen is the terminal electron acceptor,
  combining with electrons and H ions to produce
  water
• As NADH delivers more H and electrons into the
  ETS, the proton gradient increases, with H
  building up outside the inner mitochondrial
  membrane, and OH- inside the membrane.

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Proton-motive force   1. Two components to H
gradient across inner mitochondrial
membrane.   a. pH gradient (chemical
gradient)   b. voltage gradient (electrical
gradient)   Therefore, this is an electrochemical
gradient.  
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Proton-motive force (??) - expression of energy
present in electrochemical gradient. - 220 mV -
voltage component 80 pH component 20  3.
Maintenance requires inner mitochondrial membrane
be highly impermeable to H. - electron transport
is uncoupled from ATP formation by
2,4-dinitrophenol. - makes inner mitochondrial
membrane permeable to H.  4. ?? also drives ADP,
phosphate and Ca into matrix.
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d. ATP formation   1. chemiosmotic mechanism   -
energy stored in proton gradient drives
phosphorylation of ADP.   2. catalyzed by ATP
synthase.   3. ATP synthase (see Fig.5.22 5.28,
Karp 204 209)   a. F1 headpiece projects into
matrix. b. F0 basepiece embedded in lipid
bilayer - contains H channel.   4.
controlled movement of H through channel
induces   a. conformation changes   b. these
drive ATP formation.
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e. Overall products of electron transport chain
  1. ATP   - each NADH 3 ATP   - each FADH2
2 ATP   2. H2O   - formed by O2 finally
accepting electrons.
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Photosynthesis and the Chloroplast
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Heterotrophs vs Autotrophs   1.
Heterotrophs   - depend on an external source of
organic compounds.   - earliest life forms must
have been heterotrophs.   - earliest life forms
would have utilized organic molecules that had
formed abiotically. 2. Autotrophs   - utilize
CO2 to manufacture their organic molecules.  a.
Chemoautotrophs - utilize the energy stored in
inorganic molecules (e.g. hydrogen sulfide) to
convert CO2 into organic compounds.  b.
Photoautotrophs  - utilize the radiant energy of
sun to convert CO2 into organic compounds.
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Photoautotrophs include   1. higher plants 2.
eukaryotic algae 3. various flagellated
protisists. 4. a variety of prokaryotes (eg.
blue-green bacteria)   - all of the above carry
out photosynthesis   3. Photosynthesis   -
sunlight is transformed into chemical energy and
used to form carbohydrates.     - photosynthesis
in higher plants will be examined.
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So, What is this photosynthesis thing?

• It captures light energy via pigment molecules
• Chlorophylls and accessory pigments (carotenes
  and phycobilins)
• Its a redox reaction
• Produces oxidizing power (O2) via photolysis
• Captures electrons via cytochromes (plant version
  of ETC proteins) and produces reducing power in
  form of NADPH (like NADH, but without the P!(

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So, What is this photosynthesis thing?

• It produces ATP via photophosphorylation
• Couples e- transfer to proton gradients and ATP
  synthase
• It reduces CO2 to H2O
• 6 CO2 12 H2O ? C6H12O6 6 H2O 6 O2
• CO2 2 H2A ? CH2O H2O 2A

Electron Donor
Oxidized Donor
Source Carbon
Organic Carbon
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There are Two Fundamental Mechanisms in
Photosynthesis

•   LIGHT Reactions - photochemical reactions
• molecular excitation of chlorophyll by light
  results in a charge () separation across a
  membrane, with generation of proton motive force
  ( H gradient), reduction of NADP via an ETC to
  NADPH
• DARK Reactions - thermochemical reactions carbon
  dioxide reduction (fixation) -  occurs in 3
  stages carboxylation   CO2   RuBP (5c)  --gt   
  2 PGA (3c) reduction  of  PGA    NADPH  --gt   
  PGAL regeneration of  RuBP  via  HMP  pathway

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Light Reaction

•  
• Energy from sunlight is converted to chemical
  energy
• Products
• i. ATP
• ii. NADPH
• Occurs in thylakoid membranes.
•  

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Dark Reaction

• ATP and NADPH are used to synthesize
  carbohydrates
• Occurs in stroma

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The Chloroplast   - organelle in which
photosynthesis takes place.   - located
predominantly in mesophyll cells of leaves.   -
semi-autonomous and self replicating.   - bounded
by 2 membranes separated by a narrow space.   -
thylakoids - flattened membranous sacs within
chloroplast.   a. lumen - space inside a
thylakoid   b. grana - orderly stacks of
thylakoids.   - stroma - space surrounding
thylakoids.
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• 2. Absorption of light
•  
• Energy comes from sun in form of electromagnetic
  radiation
• This radiation travels in discrete packets called
  photons
• When photon is absorbed, compound is converted to
  a higher-energy state (excited state).
•  - Ground state may be re-established in three
  different ways
• 1. Energy may be dissipated as heat.
• 2. Energy may be re-emitted in form of longer
  wavelength (fluorescence).
• 3. Energy may be transferred to another molecule.
  
• This is what happens with photosynthetic
  pigments.

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chloroplasts
library.thinkquest.org/ 3564/Cells/cell120.gif
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3. Photosynthetic Pigments   - pigments are
molecules that contain a chromophore.   -
chromophore chemical group capable of absorbing
light of particular wavelengths (?).   -
absorption spectrum plot of intensity of light
absorbed vs ?.   - action spectrum plot of
physiological response vs ?.   - action spectrum
of photosynthesis follows absorption spectrum of
the chlorophylls and carotenoids.
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• Chlorophylls
• major light-capturing molecules
•  
• - absorb light of blue and red ?  
• - reflect green ? (we see)
•  
• - two parts
• a. porphyrin ring functions in light
  absorption
• - magnesium atom in center of ring
• - different side groups on ring.
• - this gives different kinds of chlorophyll (a
  b c d)
•  
• b. phytol side-chain
• - inserts chlorophyll in lipid bilayer of
  thylakoid
• membrane.

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Chlorophyll
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b. Carotenoids an accessory pigment - long
hydrocarbon chains containing alternating double
bonds.   - increase efficiency by absorbing light
in those regions where chlorophyll absorbs light
inefficiently. - absorb light of blue and green
? (400-550nm) . - reflect yellow, orange and
red ? (gt550 nm, lt700 nm). - protect
photosynthetic machinery from damage caused by
reactive oxygen species.
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Carotenoid Structure
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4. Reaction-center chlorophyll   - specific
chlorophylls capable of transferring electrons to
an electron acceptor.   - other pigments act as a
light-harvesting antenna system.   - absorb light
at other ? and transfer energy to reaction center
chlorophyll.   - reaction-center chlorophylls are
organized into 2 photosystems.   - these linked
by a chain of electon carriers.   - all within
thylakoid membranes.
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Photosystem II (PSII) 1. Reaction center
chlorophyll is referred to as P680.   P
pigment 680 ? of light that this
chlorophyll molecule absorbs most
strongly.    2. When P680 absorbs photons, it
gives up electrons to a primary electron acceptor
(pheophytin) of higher reducing potential. 3.
P680 replenishes its electrons from H2O. 4. as
H2O becomes oxidized, O2 is released. -
Pheophytin eventually transfers electrons to a
chain of electron carriers.
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Photosystem I (PSI)   1. Reaction center
chlorophyll is P700.   2. P700 accepts
electrons from last member of electron transport
chain of PSII. 3. P700 is raised to an excited
state by absorbing light. 4. In this state
electrons are transferred to primary electron
acceptor Ao. 5. Ao.has a high reducing
potential. Electrons have two fates. They can
pass down  a. a short electron transport chain
to NADP to form NADPH.   b. to P700 to form
ATP (cyclic photophosphorylation).
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5. Z scheme or pathway
1. two photosystems acting in series.   2.
electron flow occurs in 3 steps   a. between H2O
and PSII.   b. between PSII and PSI. Electron
transport chain (see Fig. 6.16 but not all
details)   c. between PSI and NADP.   3. as
electrons flow along Z-pathway, H ions are moved
from stroma to inner compartment of thylakoids.
  4. important end result is proton
gradient.   proton concentration   a. high in
lumen of thylakoid.   b. low in stroma.
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6. Photophosphorylation 1. formation of ATP as
result of electrons moving through photosystems I
and II. 2. as in mitochondria   a. proton
gradient drives ATP formation.   b. enzyme is
ATP synthase.   3. ATP synthase embedded in
thylakoid membranes.   4. two types of
photophosphorylation   a. noncyclic
photophosphorylation   b. cyclic
photophosphorylation
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a. Noncyclic photophosphorylation   1.
electrons move in linear path from H2O to
NADP   2. uses photosystems I and II   3.
formation of ATP, NADPH2 and O2   b. Cyclic
photophosphorylation 1. electrons move from
P700 to ferredoxin and back to P700   2. involves
photosystem I only.   3. formation of
ATP   Relative amount of noncyclic vs cyclic
photophosphorylation is regulated.
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7. CO2 fixation and formation of carbohydrate   -
done in all photosynthetic plants by C3
cycle.   C3 or Calvin cycle (Fig. 6.18, Karp, p.
241) 1. Carboxylation   a. CO2 combines with
ribulose-1, 5-bisphosphate.   b. forms a
transient 6-carbon compound.   c. this breaks
down to form 2 molecules of 3-
phosphoglycerate (PGA). d. catalyzed by
ribulose-1, 5-bisphosphate carboxylase
(Rubisco). 2. ATP is used to form 1, 3
bisphosphyglycerate . 3. NADPH is used to
reduce above to glyceraldehyde 3-phosphate (GAP)
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GAP has a number of fates 1. Remain in
chloroplast.   a. regenerate RuBP   b.
converted to starch 2. Exported to cytosol.
a. converted to sucrose.   b. oxidized in
glycolysis and TCA cycle to provide ATP