Chapter 5

Mitochondrion, Aerobic Respiration, and

Energy Generation
 Mitochondria
 Structure
 Function
 Organization and distribution
 Mitochondria DNA
 Aerobic Respiration

http://www.ncbi.nlm.nih.gov/books/NBK26894/
http://en.wikipedia.org/wiki/Mitochondrion
 The Mitochondria, Mit
Introduction
 The mitochondrion (plural, mitochondria) is a membrane-bound
organelle found in most eukaryotic cells.
 Mits are described as "the powerplant of the cell" because they
generate most of the cell's supply of adenosine triphosphate
(ATP), used as a source of chemical energy.
 Mits are also involved in cell signaling, cellular
differentiation, cell death, as well as maintaining the control
of the cell cycle and cell growth.
 Mits have been implicated in several human diseases,
including mitochondrial disorders, cardiac dysfunction,
heart failure and even autism ( 自闭症 ).
http://www.ncbi.nlm.nih.gov/pubmed/21119085/
 Several unique characteristics of Mit
 The number of Mit in a cell can vary widely by organism,
tissue, and cell type. For example, RBCs have no Mit,
whereas liver cells can have more than 2000. The size and
number of Mit reflects the energy requirements of the cell.
 Mit is composed of compartments that carry out specialized
functions. These compartments or regions include the outer
membrane, the intermembrane space, the inner membrane,
the cristae ( 线粒体嵴 ) and matrix ( 基质 ).
 The Mit plasticity is dynamically regulated. Mit proteins
vary depending on the tissue and the species. For example,
there are 615 distinct proteins identified from human
cardiac mitochondria, whereas 940 proteins found in rat.
 The Mit has its own independent genome. Further, its DNA
shows substantial similarity to bacterial genomes.
 The size and number of
mitochondria reflect the energy
requirements of the cell (a), a
Isolated Mit living fibroblast; (b), TEM of a
Normal RBCs from liver cells thin section through a Mit; (c),
localization of Mit in the mid-
piece of the sperm tail.
Localization of mitochondria near sites of high ATP utilization in
cardiac muscle and a sperm tail
 Mito-plasticity
Rapid changes of shape are observed when a
Mit is followed in a living cell.
Time-lapse micro-cinematography of
living cells shows that Mit are
remarkably mobile and plastic
organelles, constantly changing their
shape and even fusing with one another
and then separating again.
(A) 3D reconstruction of a Mit
network (green) in a yeast cell
with fluorescently labeled Mit, as
imaged by confocal microscopy.
(B) Fluorescently labeled
mammalian Mit network in a
Mitochondrial Networks: cultured fibroblast cell from an
Fusion and Division African green monkey.
The mitochondrial genome
Total 37 genes encode 2 rRNAs, 22 tRNAs, and 13 proteins.
  Structure properties of Mit
 Double-membraned Mit forms 5 compartments
 The two membranes have different properties
 Mit-associated ER membrane (MAM) is another structural
element that is increasingly recognized for its critical role in
cellular physiology and homeostasis.

Basic elements of Mit

 The outer membrane
 The intermembrane space
 The inner membrane
 The cristae space
 The Matrix
Outer membrane  It encloses the entire organelle.
 It is about 60 to 75 Å thick.
 Protein: phospholipid is about 1:1.
 Contains a large number of Porins.
 Permeable to molecules of 5kD or
less.
It also contains enzymes involved in
such diverse activities as the elongation
of fatty acids, oxidation of epinephrine,
and the degradation of tryptophan.
These enzymes include monoamine
oxidase (MAO), cytochrome c-
reductase, and fatty acid Co-A ligase.

How can larger proteins enter the Mit through outer membrane?
Porin
Intermembrane space  It is the space between the
outer and the inner
Schematic diagram of the Mit
membrane, also known as
perimitochondrial space.
 The concentrations of small
molecules (?) in this space is
the same as the cytosol.
 One marker protein in this
space is cytochrome c.
 It contains several enzymes
use ATP to phosphorylate
other nucleotides.

How can the intermembrane space maintain its chemical equivalent to

the cytosol with respect to the small molecules it contains?
Inner membrane is the  The inner membrane surrounds
the matrix of Mit and is highly
major working part of Mit specialized.
 It contains the “double”
phospholipid cardiolipin.
 It doesn't contain porins, but
contains a variety of transport
proteins that make it highly
impermeable to all molecules.
 It has a very high protein-to-
phospholipid ratio (more than 3:1)
Schematic diagram of the  It contains proteins with five types
Inner membrane and cristae of functions:
① Electron-transport chain: Carry out oxidation reactions
② ATP synthase: Makes ATP in the matrix
③ Transport proteins: Allow the passage of metabolites
④ Protein import machinery, enter and localization in Mit
⑤ Mitochondrial fusion and fission protein
Cristae is a special structure of Mit, which
compartmentalized by a series of inner Mit
membrane infoldings.

 It expands the surface area of the

inner Mit membrane, enhancing
its ability to produce ATP.
 The area of the inner membrane
is about five times as large as the
outer membrane.
 The folds of cristae are studded
with small round bodies known as
F1 particles or oxysomes.
Schematic diagram of the
Inner membrane and cristae
Matrix is the space enclosed
 It contains about 2/3 of the total
by the inner membrane.
protein in a Mit.
 It contains a highly
concentrated and selected set of
mixture of hundreds of
enzymes and their substrates.
 The matrix is important in the
production of ATP with the aid
of the ATP synthase contained
in the inner membrane.
 Mitochondria have their own
genetic material, and the
machinery to manufacture their
own RNAs and proteins.
Protocol for isolation the Outer Membrane, Inner Membrane,
and the Two Internal Compartments of Mit
Biochemical fractionation of purified
mitochondria into separate components
http://www.ncbi.nlm.nih.gov/pubmed/17406588
These techniques have made
it possible to study the
different proteins in each
mitochondrial compartment.
The method shown allows the
processing of large numbers
of mitochondria at the same
time. It takes advantage of the
fact that, in a solution of low
osmotic strength, water flows
into mitochondria and greatly
expands the matrix space
(yellow). While the cristae of
the inner membrane unfold to
accommodate the expansion,
the outer membrane—which
has no folds—breaks,
releasing a structure
composed of only the inner
membrane and the matrix
Localization of metabolic functions within the mitochondrion
Outer membrane:
Phospholipid synthesis
Fatty acid desaturation
Fatty acid elongation
Marker: monoamine oxidase
(MAO), 单胺氧化酶
Matrix
Pyruvate oxidation
TCA cycle
ß oxidation of fats
DNA replication, RNA
transcription,
Protein translation
Marker: MDH (malate
dehydrogenase), 苹果酸脱氢酶
Inner membrane and critae:
Electron transport
Oxidative phosphorylation
Metabolite transport
Cardiolipin/TPL(20%)
Marker: cytochrome oxidase, 细
胞色素氧化酶
Intermembrane space
Nucleotide phosphorylation
Marker: adenylate kinase,
腺苷酸激酶
 Functions of Mit
 Energy conversion, the most dominant role for the
mitochondria is the production of ATP.
 Pyruvate and the citric acid cycle, the acetyl-CoA is the
primary substrate to enter tricarboxylic acid (TCA) cycle.
 NADH and FADH2: the electron transport chain, the redox
energy from NADH and FADH2 is transferred to oxygen in
several steps via the electron transport chain to produce ATP.
 Heat production
 Storage of calcium ions
 Additional functions including signaling through ROS,
regulation of the membrane potential, apoptosis, calcium
signaling, regulation of cellular metabolism, steroid synthesis,
and hormonal signaling etc.
An overview of carbohydrate metabolism in eukaryotic cells
Organelle DNA and protein importing
A. Organelle DNA
The size range of organelle DNA is similar to that of viral DNAs.
 Mit DNAs range from 6000 ～ 300000bp (some land plants). DNA of Mit
genome(in mammals) is about 16500bp (<0.001% of nuclear genome). Chl
genomes are about 10 times larger than Mit and contain about 120 genes.
 Chl DNA: from 70000to 200000bp (genome of land plants);
 Mit has its own genetic systems
 Genes in mtDNA encode rRNAs, tRNAs, and some
mitochondrial proteins.
Human mt DNA: 16,569bp
2 rRNAs （ 16s and 12s RNA ） , 22 tRNAs,
13 polypeptides: NADH reductase. 7 sub.
Cyt b-c1 complex. 1 cytb
Products of mt genes
are not exported Cyt oxidase. 3 subunits
ATP synthase: 2 F0 sub
Mit and Chl are
organelles
semiautocephaly.
The synthesis of
mt proteins is
coordinated
Aerobic Respiration and Energy Generation

Oxidation of
carbohydrate:
Glucose
↓ glycolysis in cytosol
Pyruvate and NADH
↓ decarboxylation
Acetyl CoA
↓ TCA cycle
2CO2+FADH2+3NADH+
3H++GTP+HS-CoA

Decarboxylated by the dehydrogenase

Molecular basis of
oxidative phosphorylation
A. Molecular basis of oxidation: Electron-
transport chain

NADH, Nicotinamide
adenine dinucleotide
FADH2, reduced flavin
adenine dinucleotide
B. Molecular basis of phosphorylation:
ATP synthase
 The structure of the ATP synthase
 F1 particle is the catalytic subunit, which contains
5 types of subunits in the ratio of 3: 3: 1: 1: 1.
 F0 particle attaches to F1, which contains 3 types of
subunits in the ratio of 1a ： 2b ： 12c, and is
embedded in the inner membrane.

F1

F0
 The ATP synthase is a reversible coupling device
 Proton translocation through F0 drives ATP synthesis
by F1: Binding Change Model and Rotational Catalysis

Boyer proposed in
1979, and was
greatly stimulated
by the publication
in 1994 of the
structure for F1
complex (X-ray)
from bovine heart
mitochondria

结合变换机制
旋转催化模型
C. Mithchell’s Chemiosmotic theory (1961)
 The pH and electrical gradient resulting from
transport of protons links oxidation to
phosphorylation.
 When electrons are passed to carriers only able to
accept electrons, the H+ is translocated across the
inner membrane.

More than 21026 molecules (>160kg) of ATP per day in our bodies.
 Electrons pass from NADH or FADH2 to O2, the terminal electron
acceptor, through a chain of carriers in the inner membrane (FMN,
Fe-S center, Heme group Fe, CoQ).
 As electrons move through the electron-transport chain, H+ are
pumped out across the inner membrane, and form Proton motive
force.

 Electrons move through the inner membrane via a

series of carriers of decreasing redox potential.
A testable prediction followed
from Mitchell’s hypothesis

If not all the

detergent is
removed,
what will
happen?
Summary of the major activities during aerobic
respiration in a mitochondrion

NADHO2: 3ATP/2e;
FADH2 O2 : 2ATP/2e
 The proliferation and origin of Mit and Chl.
A. Organelle growth and division determine the
number of Mitochondria and Plastids in a cell

Mit fission and fusion (a dividing Mit in a liver cell);

Dividing or Budding of Mit.
Chloroplasts: dividing and formation of chloroplasts from
proplastids begins by the light-induced budding of the inner
membrane.
B. Origin: The
endosymbiont theory
( 内共生学说 )

Compare the
ribosomal RNA with
the base sequence of
various bacterial
rRNAs:
Purple bactria-
Mitochondria
Cyanobacteria-
Chloroplasts