Lab 8

1. December
13, 2015
PRACTICAL CELL BIOLOGY LAB 8
Mitochondria
Introduction
Mitochondria called the “powerhouse” of the cell. While metaphorically
correct, biochemical events occurring in this tiny organelle. For instance, it is now
well known that these events are not only producing energy but also playing
intriguing roles for immunity and cell death. The energy produced by
mitochondria, is in the form of a molecule called adenosine triphosphate (ATP),
which effectively stores energy in its chemical bonds between phosphate
molecules. Making and breaking those bonds stores and releases energy,
respectively. The ATP formed by mitochondria can be distributed throughout the
cell to facilitate other chemical processes.
Past and current mitochondrial research has been performed on
mitochondria prepared from rat liver, rat heart, or beef heart. Toward the end of the
1980s, a new branch of human pathology began with the discovery of human
disorders linked to mitochondrial dysfunction. Therefore, it became necessary to
investigate and to study the status of mitochondria in human tissue. Actually we
have to know that it is not always possible to obtain large amounts of human
tissues for extracting mitochondria.
The energy required to power living cells comes from the sun. Plants
capture this energy and convert it to organic molecules. Animals in turn, can gain
this energy by eating plants or other animals. The energy that powers our cells is
obtained from the foods we eat. The most efficient way for cells to harvest energy
stored in food is through cellular respiration, a catabolic pathway for the
production of adenosine triphosphate (ATP). Glucose, derived from food, is broken
down during cellular respiration to provide energy in the form of ATP and heat.

2. December
13, 2015
PRACTICAL CELL BIOLOGY LAB 8
Cellular respiration has three main stages: glycolysis, the citric acid cycle and
electron transport chain. The Krebs cycle (citric acid cycle) and electron
transport occur within mitochondria, so that the mitochondria are known as the
powerhouses of the cell.
In general, the number of mitochondria per cell and the number of
cristae per mitochondrion are related to the energy requirement for the function
carried out by that cell type. So the number of mitochondria in a cell depends on
the cell's function. Cells with particularly heavy energy demands, such as muscle
cells, have more mitochondria than other cells. That is mean a cell rich in
mitochondria is likely to be a very energetic cell. A cell poor in mitochondria is
less energetic such as adipose cell.
Origin of mitochondria
Mitochondria show many similarities to free-living procaryotic organisms,
for example, they often resemble bacteria in size and shape, they contain DNA,
they make protein, and they reproduce by dividing in into two.
Structure of mitochondria
The outer membrane of mitochondria is smooth and separates the
mitochondrion from the cytoplasm. In contrast, the inner membrane has
numerous invaginations called cristae, that is mean it forms septum-like
folds (cristae), which extend to various lengths across the organelle. These
membranes divides mitochondrion into two submitochondrial
compartments: the intermembrane space between the outer membrane
and the inner membrane with its cristae, which it is the narrow liquid part
between the two membranes, separated by about 8nm, and the matrix,

3. December
13, 2015
PRACTICAL CELL BIOLOGY LAB 8
which it is the part enclosed by the innermost membrane. Several of the
steps in cellular respiration occur in the matrix due to its high
concentration of enzymes, also matrix contains mitochondrial
deoxyribonucleic acid (mtDNA) and mitochondria reproduce
independently of the cell by fission.
Size and shape of the mitochondria
Size and shapes of mitochondria can be very different. On average, they are
0. 25μm thick and 2–7μm long. There are also giant forms (giant mitochondria).
The number of cristae mitochondriales also varies. As a result, the sizes of the
inner and outer compartments are also different. The folds that originate at the
inner membranes and extend into the inner centers of the mitochondria—the cristae
mitochondriales—are different in length and form a series of incomplete transverse
septa.

4. December
13, 2015
PRACTICAL CELL BIOLOGY LAB 8
The mitochondrion is composed of two layers of phospholipids making an
outer smooth membrane that forms a characteristic ball- or bean-shaped structure
and a highly folded inner membrane. The inner membrane marks the location for
the biochemical events (the electron transport system), which will produce
energy for the cell. By folding the inner membrane, the surface area is increased
two to three times compared to the flat surface area, and there by more energy
can be produced.
Mitochondria and energy production
Most eukaryotic cells contain many mitochondria, which occupy up to 25
percent of the volume of the cytoplasm. These complex organelles, the main sites
of ATP production during aerobic metabolism. The two membranes that bound a
mitochondrion differ in composition and function. The outer membrane, composed
of about half lipid and half protein. The inner membrane, which is much less
permeable, is about 20 percent lipid and 80 percent protein. In
nonphotosynthetic cells, the principal fuels for ATP synthesis are fatty acids and
glucose. The complete aerobic degradation of glucose to CO2 and H2O is coupled
to the synthesis of as many molecules of ATP. In eukaryotic cells, the initial
stages of glucose degradation take place in the cytosol, where 2 ATP molecules
per glucose molecule are generated. The terminal stages of oxidation and the
coupled synthesis of ATP are carried out by enzymes in the mitochondrial matrix
and inner membrane. so many ATP molecules per glucose molecule are generated
in mitochondria.

5. December
13, 2015
PRACTICAL CELL BIOLOGY LAB 8
The size, Coding Capacity and inheritance of mtDNA
Mitochondrial DNA (mtDNA ) is the DNA located
in organelles called mitochondria, surprisingly, the size of the mtDNA, the number
and nature of the proteins it encodes, and even the mitochondrial genetic code
itself varies greatly between different organisms. In humans, mitochondrial DNA
can be assessed as the smallest chromosome coding for only 37 genes and
containing only about 16,600 base pairs. Human mitochondrial DNA was the first
significant part of the human genome to be sequenced. In most species, including
humans, mtDNA is inherited solely from the mother. The mtDNA is organized as
a circular, double-stranded DNA.
In mammals, each double-stranded circular mtDNA molecule consists of
15,000-17,000 base pairs. The two strands of mtDNA encodes 37 genes, 13 are for
proteins (polypeptides), 22 are for transfer RNA (tRNA) and two are for the small
and large subunits of ribosomal RNA (rRNA).
Mitochondria contain multiple copies of their own DNA genomes, which
code for some of the mitochondrial proteins. Because each human inherits
mitochondrial DNA only from his or her mother (it comes with the egg but not the
sperm), the distinctive features of a particular mitochondrial DNA can be used to
trace maternal history.

6. December
13, 2015
PRACTICAL CELL BIOLOGY LAB 8
Mitochondria and cell death
 In a healthy cell, the outer
membranes of its
mitochondria display the
protein Bcl-2 on their
surface. Bcl-2 inhibits
apoptosis.
 Internal damage to the cell
o causes a related
protein, Bax, to migrate to the surface of the mitochondrion where it
inhibits the protective effect of Bcl-2 and inserts itself into the outer
mitochondrial membrane punching holes in it and causing
o cytochrome c to leak out.
 The released cytochromec binds to the protein Apaf-1 ("apoptotic protease
activating factor-1").
 Using the energy provided by ATP,
 These complexes aggregate to form apoptosomes.
 The apoptosomes bind to and activate caspase-9.
 Caspase-9is one of a family of over a dozen caspases.
 Caspase-9cleaves and, in so doing, activates other caspases (caspase-3and -
7).
 The activation of these "executioner" caspases creates an expanding cascade
of proteolytic activity which leads to
o digestion of structural proteins in the cytoplasm,
o degradation of chromosomalDNA, and
 phagocytosis of the cell.