III Pharm D (2011-2012), Pharmacology-II (T), PESCP, Bangalore-50, KPS Gowda Asst.Prof. 6. The dynamic cell: The structures and functions of the components of the cell a) Cell and macromolecules: Cellular classification, subcellular organelles,macromolecules, large macromolecular assemblies b) Chromosome structure: Pro and eukaryotic chromosome structures, chromatin structure, genome complexity, the flow of genetic information. c) DNA replication: General, bacterial and eukaryotic DNA replication. d) The cell cycle: Restriction point, cell cycle regulators and modifiers. e) Cell signaling: Communication between cells and their environment, ionchannels, signal transduction pathways (MAP kinase, P38 kinase, JNK, Ras and PI3-kinase pathways, biosensors. a)Cell and macromolecules: Cellular classification, subcellular organelles, macromolecules, large macromolecular assemblies- The cell comes from the Latin cellula, meaning “a small room”. The cell is the structural and functional unit of all known living organisms. The cell was discovered by Robert Hooke in 1665. Human contains about 10 trillion cells. Most plant and animal cells are between 1 and 100μm and therefore are visible only under the microscope. Types of cell: There are two types of cell. Prokaryotic and eukaryotic. Prokaryotic cells: Prokaryotic word derived from Greek meaning- before nuclei. Examples- cells in the bacteria and cyano bacteria (blue green algae).These cells have few internal structures. They do not have membrane bound nucleus. The bacterial cells are very small (about 1-2 μm diameter and 10 μm long). These cells have 3 shapes- rod, spherical and spiral. The cell division is by binary fission. The structural components of prokaryotic cells: The nuclear material of prokaryotic cell consists of a single chromosome. The flagella and pili are projected from the cell surface. These consist of proteins. They facilitate the movement and communication between the cells. The cells enclosed by cell envelop. This consists of plasma membrane and cell wall. Some bacteria
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III Pharm D (2011-2012), Pharmacology-II (T), PESCP, Bangalore-50, KPS Gowda
Asst.Prof.
6. The dynamic cell: The structures and functions of the components of the cell
a) Cell and macromolecules: Cellular classification, subcellular organelles,macromolecules,
large macromolecular assemblies b) Chromosome structure: Pro and eukaryotic chromosome
structures, chromatin structure, genome complexity, the flow of genetic information. c) DNA
replication: General, bacterial and eukaryotic DNA replication. d) The cell cycle: Restriction
point, cell cycle regulators and modifiers. e) Cell signaling: Communication between cells and
their environment, ionchannels, signal transduction pathways (MAP kinase, P38 kinase, JNK,
Ras and PI3-kinase pathways, biosensors.
a)Cell and macromolecules: Cellular classification, subcellular organelles, macromolecules,
large macromolecular assemblies- The cell comes from the Latin cellula, meaning “a small
room”. The cell is the structural and functional unit of all known living organisms. The cell was
discovered by Robert Hooke in 1665. Human contains about 10 trillion cells. Most plant and
animal cells are between 1 and 100µm and therefore are visible only under the microscope.
Types of cell: There are two types of cell. Prokaryotic and eukaryotic.
Prokaryotic cells: Prokaryotic word derived from Greek meaning- before nuclei. Examples-
cells in the bacteria and cyano bacteria (blue green algae).These cells have few internal
structures. They do not have membrane bound nucleus. The bacterial cells are very small (about
1-2 µm diameter and 10 µm long). These cells have 3 shapes- rod, spherical and spiral. The cell
division is by binary fission.
The structural components of prokaryotic cells: The nuclear material of prokaryotic cell
consists of a single chromosome. The flagella and pili are projected from the cell surface. These
consist of proteins. They facilitate the movement and communication between the cells. The
cells enclosed by cell envelop. This consists of plasma membrane and cell wall. Some bacteria
also have another layer- capsule. This envelop gives the rigidity to the cell. Inside the cell is the
cytoplasmic region that contains the cell genome (DNA), ribosomes and various cell inclusions.
The DNA is condensed to a nucleiod. There are circular structures called plasmids, which carry
extrachromosomal DNA.
2. Eukaryotic cells: Eukaryotic word is derived from Greek- eu means good and karyon means
nut or kernel. These cells contain complex structures enclosed within the membranes. The
membrane bound nucleus is present in these cells. Most of these cells also contain other
membrane bound organelles such as mitochondria, chloroplast and the Golgi apparatus. All
species of large complex organisms are eukaryotes, including animals, plants and fungi.
Organelles of the eukaryotic cells: 1.Lysosomes: These are cellular organelles that contain
acid hydrolase enzymes to break down waste materials and cellular debris. They are found in
animal cells. The membrane around a lysosome allows the digestive enzymes to work at the
acidic pH. The lysosomes are formed from the Golgi apparatus. The aged cell organelles are
degraded in a lysosome is called autophagy. They are also called as suicide-bags or suicide-sacs
due to their autolysis. The size of the lysosomes varies from 0.1 to 1.2µm. The pH of the interior
of the lysosomes is 4.8 and it is acidic compared to the slightly alkaline cytosol (pH 7.2). The
lysosome maintains this pH differential by pumping protons (H+ ions) from the cytosol across
the membrane via proton pumps and chloride ion channels. The lysosomal membrane protects
the cytosol, and rest of the cell, from the degradative enzymes of the lysosome. If lysosomal acid
hydrolases of lysosomes leaked into cytosol , these enzymes fails to produce their effects in the
alkaline environment. Some of the examples of digestion by the acid hydrolase enzymes.-
Nuclease degrade RNA and DNA into their mononucleotides building blocks. Protease degrades
proteins to peptides. Phosphatases remove phosphate groups from mononucleotides,
phospholipids and other compounds.
Tay-Sachs disease is caused by the deficiency of lysosomal enzyme that digests gangliosides (a
glycolipid). This results in the accumulation of these glycolipids in the neurons. It is inherited
disease; the affected children commonly become demented and blind by age 2 and die before
their 3rd
birthday.
Autophagy- It is a catabolic process involving degradation of a cell’s own components through
lysosomal enzymes. It involves the formation of a membrane around a targeted region of the cell,
separating the contents from the rest of the cytoplasma. The resultant vesicle fuses with a lysome
and subsequently degrades the contents.
Autolysis- It is the spontaneous disintegration of cells or tissues by lysosomal enzymes, as
occurs after death and in some pathological conditions.
Peroxisomes: These are roughly spherical organelles. All animal cells (except RBC) contain
peroxisomes. Its diameter is 0.2 to 1µm.Peroxisomes contain several oxidases-enzymes that use
molecular oxygen to oxidize organic substances, in the process forming H2O2. Peroxisome also
contains catalase, which degrades hydrogen peroxide to yield water and oxygen.
2H2O2 -------- catalase2H2O +O2
In X-linked adrenoleukodystrophy (ADL), the peroxisomal oxidation of very long chain fatty
acids is defective. This damages the white matter of the brain and impairs the adrenal glands.
3.Endoplasmic reticulum: It is an extensive network of closed, flattened membrane-bounded
sacs called cisternae. In smooth ER the synthesis of fatty acids and phospholipids takes place.
The hepatocytes have abudant smooth ER. Enzymes in the smooth ER of the liver also modify or
detoxify hydrophobic chemicals such as pesticides and carcinogens into more water soluble,
conjugated products that can be excreted from the body. The smooth ER is one of the sites of
drug metabolism by cytochrome-450 enzymes. High doses of such compounds result in
enlargement of smooth ER in the liver cells.
The proteins manufacturing ribosome are present on the surface of the rough ER. The ribosome
are bind with the riboporin (a glycoprotein receptor) expressed on the surface of the RER. These
receptors are not present on the smooth ER
The functions of RER – It facilitates the folding of newly synthesized proteins.Only properly
folded proteins are transported from RER to Golgi complex.RER helps for the transportation of
newly synthesized proteins. The RER helps for the insertion of proteins into the endoplasmic
reticulum membrane. It helps for glycosylation- attachment of oligosaccharides to a protein. It
helps for disulfide bond formation, as this helps for the stabilization of tertiary and quaternary
structure of the proteins.
4.The Golgi complex- It is the cell organelle of the eukayotic cells. It was identified by Italian
Physician Camello Golgi in 1898. The stack of Golgi cisternae has 3 regions- the cis, the medial,
and the trans. Transport vesicles from the RER fuse with the cis region of the Golgi complex,
where they deposit their protein contents. These proteins then progress from the cis to the medial
and then to the trans region. Within each region different enzymes modify the proteins. The
modified proteins get packed into the secretory vesicles. The functions of Golgi complex are
1.Enzymes present in the cisternae modify proteins- glycosylation,phosphorylation,etc. 2.
Transportation of lipids within the cell. 3. Formation of lysosomes. 4. Play a role in the synthesis
of the proteoglycons and carbohydrates.
5.Mitochondria (singular-mitochondrion)- Mitochondrion is derived from–mitos meaning
thread, chondrion- granule. These are the membrane-enclosed organelles found in the eukaryotic
cells. Size - 0.5 to 1 micrometer (µm) in diameter. These are described as power houses of the
cell, as they generate ATP. The two membranes that bound a mitochondrion differ in
composition and function. The outer membrane composed of about half lipids and half proteins.
This is permeable to molecules having molecular weight as high as 10,000. The inner membrane
is less permeable, is composed of 20% lipids and 80% proteins. The large number of in folding
of the inner membrane (cristae) increases the surface area. The ATPs are synthesized from fatty
acids and glucose. The complete aerobic degradation of glucose to CO2 and H2O leads to 30
molecules of ATP. In eukaryotic cells, the initial stages of glucose degradation takes place in the
cytosol, where 2 ATP molecules per glucose are generated. The terminal stages of oxidation and
synthesis of ATP are carried out by enzymes in the mitochondrial matrix and inner membrane.
28 ATP molecules per glucose molecule are generated in mitochondria.
6.Cytoskeleton: The eukaryotic cells contain 3 kinds of cytoskeletal structures, which are
microfilaments, intermediate filaments and microtubules.
a.Microfilaments: (actin filaments)- These are solid rods about 7nm in diameter. They are also
called actin filaments, because they are built from molecules called actin (a protein). A
microfilament is a twisted double chain of actin subunits. Microfilaments are present in all
eukaryotic-
cells.
b.Intermediate filaments. Its average size is 10nm. It is mainly present in the cytoplasm. But
lamin intermediate filament found in the nucleus.
c.Microtubules: Hallow tube like structures, 24nm diameter.
Each type of cytoskeletal filament is a polymer of protein subunits. Monomeric actin subunits
assemble into microfilaments, dimeric subunits composed of α and β-tubulin polymerize into
microtubules. The intermediate filaments are composed of different proteins, examples- lamins,
keratin, etc. The cytoskeleton provides shape to the cell and helps for the movement the cell
organells and cell.
7.Nucleus: It is the largest cell organelle in animal cells, is surrounded by two membrane made
up of phospholipids and proteins. The two nuclear membranes appear to fuse at nuclear pores,
through which material moves between the nucleus and the cytosol. In a growing or
differentiating cell, the nucleus is metabolically active, replicating DNA and synthesizing rRNA,
tRNA, and mRNA. Most of the ribosomal RNA is synthesized in the nucleolus. The DNA is
packaged into chromosomes. In a nucleus that is not dividing, the chromosomes are dispersed.
During cell division the individual chromosomes are visible by light microscope.
Macromolecules: Organic molecules are molecules that contain carbon and hydrogen. All living
things contain these organic molecules. Small organic molecules can combine into very large
molecules that are called macromolecules. Macromolecules are usually polymers (poly= many,
mers= parts). A polymer is a large molecule formed by the covalent bonding of many identical or
similar small building-block molecules called monomers. Usually the reaction that joins two
monomers is a dehydration synthesis. In this type of reaction, hydrogen is removed from one
monomer and a hydroxyl group is removed from other to form a water molecule.
Macromolecules such as carbohydrates, lipids, proteins, and nucleic-acids are assembled in cells
via dehydration synthesis reactions. There are four basic kinds of biological macromolecules.
They are carbohydrates, lipids, proteins and nucleic acids. The polymers are composed of
different monomers and serve different functions.
Carbon
Carbon has four electrons in its outer shell.
Hydrogen has one electron and one proton.
Carbon can bond by covalent bonds with as many as 4 other atoms.
Methane molecule.
Carbon also form double covalent or triple covalent bonds..
Carbon can form 4 covalent bonds because it has 4 electrons in its outer shell. It can form the
following number of bonds..
4 single bonds
2 double bonds
1 double bond and 2 single bonds
1 triple bond and 1 single bond
Long chains of carbon are common. The chains may be branched or form
rings.
Hydrophilic and hydrophobic:
Polar and ionic molecules have positive and negative charges and are therefore attracted to water
molecules because water molecules are also polar. They are said to be hydrophilic because they
interact with (dissolve in ) water by forming hydrogen bonds.
Non polar molecules are hydrophobic (means “water fearing”). They do not dissolve in water.
Non polar molecules are hydrophobic.
Polar and ionic molecules are hydrophilic.
Functional groups
Organic molecules may have functional groups attached. Example, COOH is a carboxyl group.
The letter R is used to indicate an organic molecule. For example, the diagram below can
represent a carboxylic group. The R can be any organic molecule.
If polar or ionizing functional groups are attached to hydrophobic molecules, the molecule may
become hydrophilic due to the functional group. Some ionizing functional groups are:- COOH,
OH, CO and NH2. Some important functional groups are shown below.
Name
Structure
Non-ionized Ionized
Hydroxyl
Carboxyl
Amino
Phosphate
Sulfhydryl
Aldehyde
Ketone
Isomers - Different molecules that are composed of the same number and kinds of atoms are
called isomers. Glucose and fructose are both C6H12O6 but the atoms are arranged differently in
each molecule.
Structural isomers differ in their overall construction as shown above for glucose and fructose.
Geometric isomers maintain the same carbon skeleton but a double bond occurs between carbon
atoms. The two molecules below are geometric isomers because the double bond cannot rotate. If
the bond between the two carbon atoms were a single bond, they would not be isomers because
atoms attached by single bonds can rotate.
Enantiomers are molecules that are mirror images of each other. The molecules shown below are
enantiomers.
Condensations- In order to bond the two molecules together, the first hydrogen of each molecule
is removed. This is necessary because carbon has a maximum of 4 bonds and hydrogen can have
only one.
In biological systems, macromolecules are formed by removing H from one atom and OH from
the other. The H and the OH combine to form water. Small molecules (monomers) are therefore
joined to build macromolecules by the removal of water. Example: The sugar (sucrose) can be
produced by a condensation reaction of glucose and
fructose.
Sucrose:
Hydrolysis
This is a type of reaction in which a macromolecule is broken down into small molecule. It is the
reverse of condensation.
Macromolecules and monomers: Many of the common biological (macromolecules) are
synthesized from simpler building blocks (monomers).
Example of a macromolecule monomer
Polysaccharide Monosaccharide (simple sugar)
Fat(a lipid) Glycerol, fatty acid
Protein Amino acid
Nucleic acid Nucleotide
Carbohydrates- The general formula for carbohydrates is (CH2O)n
Carbohydrates include sugars, glycogen, starches, and cellulose. They represent only 2-3% of
total body mass. In humans and animals, carbohydrates function mainly as a source of chemical
energy for generating ATP needed to drive metabolic reactions. Only few carbohydrates are used
for building structural units. Examples- Deoxyribose of DNA and ribose of RNA.
Monosaccharide and disaccharides are known as simple sugars. Oligosaccharides made up of 2-
10 and polysacharides made up of more than 10 monosaccharides.
Monosaccharides: Glucose (the main blood sugar), fructose (found in fruits), galactose (milk
sugar), deoxyribose (in DNA), ribose (in RNA). The names of most sugars end with the letters
ose
There are two isomers of the ring form of glucose. They differ in the location of the OH group on
the number 1 carbon atom.
The number 1 carbon atom of the linear form of glucose is attached to the oxygen on the number
5 carbon atom. Simple sugars store energy for cells. Cells also use simple sugars to construct
other kinds of organic molecules.(eg. Ribose and 2-deoxyribose).
Disaccharides- Disaccharides are composed of 2 monosaccharides joined together by a
condensation reaction.Examples: Sucrose (table sugar) is composed of glucose and fructose.
Like glucose, sucrose stores energy.
Lactose is found in milk. It is formed when glucose binds to galactose. The digestion of
carbohydrates involves hydrolysis reactions in which complex carbohydrates (polysaccharides)
are broken down to maltose (a disaccharide). Maltose is then further broken down to produce
two glucose molecules.
Polysaccharides-
Monosaccharides may be bonded together to form long chains called polysaccharides.
Starch and glycogen- These are polysaccharides that function to store energy. They are
composed of glucose monomers bonded together producing long chains. Animals store extra
carbohydrates as glycogen in the liver and muscles. Between meals, the liver breaks down
glycogen to glucose in order to keep the concentration of glucoses in the blood. After meals, as
glucose levels in the blood rise, it is removed from the blood and stored as glycogen. Plants
produce starch to store carbohydrates. Amylopectin is a form of starch that is very similar to
glycogen It is branched but glycogen has more branches.
Structure of glycogen or
starch
Cellulose and chitin- Cellulose and chitin are polysaccharides that function to support and
protect the organism. The cell walls of the plant composed of cellulose. The cell walls of fungi
are composed of chitin. Cellulose is composed of beta-glucose monomers; starch and glycogen
are composed of alpha-glucose.
Structure of cellulose.
The glucose monomers of chitin (N-acetyl glucosamine) have a side chain containing
nitrogen.
Cotton and wood are composed mostly of cellulose. Humans and most animals do not have
necessary enzymes to break the linkages of cellulose or chitin.
Lipids-Lipids are compounds that are insoluble in water but soluble in nonpolar solvents. Some
lipids function in long term energy storage.Lipids is also an important component of cell
membranes.Lipids make 18-25% of body mass in lean adults. Like, carbohydrates lipid contain
carbon, hydrogen, and oxygen. They do not have 2:1 ratio of hydrogen to oxygen. The
proportion of electronegative oxygen atoms in lipids is usually smaller than in carbohydrates, so
there are fewer polar covalent bonds. As a result most lipids are insoluble in polar solvents such
as water: they are hydrophobic. Lipoproteins are soluble because the proteins are on the outside
and the lipids are on inside. The lipids includes triglycerides (fats and oils),phospholipids,
steroids, eicosanoids and a variety of other lipids including fat-soluble vitamins (A,D,E and K)
and lipoproteins.
Fats and oils (Triglycerides)- A triglyceride consists of two types of building blocks, a single
glycerol molecule and three fatty acid molecules. Three fatty acids are attached by dehydration
reaction. The hydrolysis, breaks down a single molecule of triglyceride into three fatty acids and
glycerol.
Fatty acids have a long hydrocarbon (carbon and hydrogen) chain with a carboxyl (acid) group.
The chains usually contain 16 to 18 carbons. Glycerol contains 3 carbons and 3 hydroxyl groups.
It reacts with 3 fatty acids to form a triglyceride or fat molecule.
Fats are nonpolar and therefore they do not dissolve in water.
Saturated fat- These are the triglycerides containing saturated fatty acids. The saturated fatty
acids have no double bonds between carbons. Examples- Palmitic acid (C15H31 COOH), stearic
acid (C15H35COOH). Examples of saturated fats- butter, cocoa butter, coconut oil, meats, etc.
Monounsaturated fats: contain fatty acids with one double bond between two fatty acid carbon
atoms. Examples- olive oil, peanut oil, etc. These fats are thought to decrease the risk of heart
disease.
Poly unsaturated fats: contain more than one covalent bond between fatty acid carbon atoms.
Examples- corn oil, sunflower oil, soybean oil and fatty fish. These fats are believed to decrease
the risk of heart disease. Unsaturated fatty acids have at least one double bond. Each double bond
produces a “bend” in the molecule.
Molecules with many of these bends cannot be packed as closely together as straight molecules,
so these fats are less dense. As a result, triglycerides composed of unsaturated fatty acids melt at
lower temperatures than those with saturated fatty acids. For example, butter contains more
saturated fat than corn oil, and it is a solid at room temperature while corn oil is a liquid.
Phospholipids- Phospholipids have a structure like triglycerides, but contain a phosphate group
in place of the third fatty acid. The phosphate group is polar and therefore capable of interacting
with water molecule. These are amphipathic in nature, as they have both polar and nonpolar
groups.
Phospholipids form a bilayer in a watery environment. They arrange themselves so that the polar
heads are oriented toward the water and the fatty acid tails are oriented toward the inside of the
bilayer.
Steroids- Steroids differs from triglycerides. They have four rings of carbon atoms. Body cells
synthesize other steroids from cholesterol. Cholesterol has a large nonpolar region consisting of
the four rings and a hydrocarbon tail. The steroids of the body are cholesterol, estrogens,
testosterone, cortisol, bile salts, and vitamin D. These are also known as sterols because they also
have at least one hydroxyl group (-OH). Polar hydroxyl groups make sterols weakly
amphipathic.
Important functions of the steroids- 1.Cholesterol is needed for cell membrane
structure.2.Estrogen and testosterone are required for regulating sexual functions.3.Cortisol is
needed for maintaining the normal blood sugar level. 4.Bile salts are needed for lipid digestion
and absorption. 5.Vitamin D is related to bone growth.
Cholesterol Estradiol
Testosterone
Other lipids: Eicosanoids are lipids derived from a 20-carbon fatty acid called arachidonic acid.
The two subclasses of eicosanoids are the prostaglandins and the leukotries.
Other lipids also include fatty acids- which under go either hydrolysis to provide ATP or
dehydration synthesis to build triglycerides and phospholipids.
Waxes- Waxes are composed of a long-chain bonded to a long-chain alcohol. They form
protective coverings for plants and animals (plant surface, animal ears).
Proteins- Proteins are large molecules that contain hydrogen, oxygen and nitrogen. Some
proteins also contain sulfur. Proteins have complex structure. The proteins make up 12-18% of
body mass in a lean adult. Proteins have many roles in the body and are largely responsible for
Secondary structure- The secondary structure is stabilized by hydrogen bonds. There are two
types secondary structure in proteins, the α-helix and β-pleated sheet. The α-helix is held
together by hydrogen bonds between the hydrogen and oxygen atom. Examples- keratin protein
present hairs, nails and skin. In β-pleated sheet, the hydrogen bond is formed between two chains
of the amino acid chains. Example- Fibroin- spider protein.
Tertiary structure- It refers to the overall- 3-dimesional shape of the polypeptide chain. Several
bonds are present in tertiary structure. The strongest S-S covalent bond between the sulfhydryl
groups of two monomers of the aminoacid cysteine. Many weak bonds –hydrogen bonds, ionic
bonds, and hydrophobic interactions also help for the folding pattern of the proteins. These bonds
are very important in maintaining the tertiary structure of some proteins.
Quarternary structure- Some proteins contain two or more polypeptide chains that associate
to form a single protein. These proteins have quaternary structure. For example, hemoglobin
contains four polypeptide chains.
Denaturation – This occurs when the normal bonding patterns are disturbed causing the shape
of the protein to change. This can be caused by changes in temperature, pH or salt concentration.
For example, acid causes milk to curdle and heat (cooking) causes egg white to coagulate
because the proteins within them denature.
Other kinds of proteins- Simple proteins contain only amino acids. Conjugated proteins
contain other kinds of molecules. For example- glycoproteins, nucleoproteins contain nucleic
acids, and lipoprotein contains lipids.
Nucleic acids- It is a macromolecule with acid property and it was isolated from the nucleus of
cells and hence it is named as nucleic acid. It is made up of C, H, O, N and P. Nucleic acids are
found in all organisms such as plants, animals, bacteria and viruses. They are found in the
nucleus as well as in the cytoplasma. It is a long chain polymer. It is composed of monomeric
units, called nucleotides. Each nucleotide consists of a nucleoside and a phosphate group. Each
nucleoside consists of a pentose sugar and a nitrogenous base. The sugar is ribose in case of
RNA and deoxyribose in case of DNA.
The nitrogenous bases are of two types, namely purine and pyrimidine. There are two main
purine bases, adenine and guanine. There are three pyrimidine bases. They are cytosine, thymine
and uracil. Cytosine and thymine are found in DNA. Cytosine and uracil are found in RNA.
Nucleosides: A base combined with a sugar molecule is called a nucleoside. In DNA four
nucleosides are present. They are adenosine, guanosine, cytidine and thymidine. In RNA
deoxyribose is replaced by ribose and the base thymine is replaced by uracil.
Nucleotides: A nucleotide is derived from a nucleoside by the addition of a molecule of
phosphoric acid. The DNA contains four different types of nucleotides. They are adenylic acid,
guanylic acid, cytidylic acid and thymidylic acid. The RNA contains uridylic acid instead of
thymidylic acid.
Polynucleotide: a number of nucleotide units linked with one another to form a polynucleotide
chain or nucleic acid.
Nucleic acids are classified into DNA and RNA. The RNA further classified into mRNA, tRNA
and rRNA.
Deoxyribonucleic acid: (DNA): It is the molecule of heredity. It functions as the genes.DNA is
present in all cells except plant virus. In eukaryotic cells, DNA is present in the chromosomes of
nucleus. In addition, the mitochondria and plastids contain DNA. In eukaryotic nucleus, the
DNA is in the form of a double helix. In bacteria, mitochondria and plastids the DNA molecules
are circular. In viruses and bacteriophages they are coiled. The number of DNA molecules in
eukaryotic cells corresponds to the number of chromosomes per cell. DNA is made up of 3
chemical components, namely 1.Sugar, 2.Phospharic acid and 3. Nitrogenous bases.
1.Sugar:
The sugar present in the DNA is called deoxyribose. It is a pentose sugar which contains five
carbon atoms (C5H10O4). It contains one O atom less than the ribose sugar. At carbon No.2 of
deoxyribose, is present a H-C-H group. But in ribose sugar the second carbon atom contains H-
C-OH group.
2.Phosphoric acid: (H3PO4)-
Deoxyribose sugar molecule linked with one phosphate group at 5th
position and another
phosphate group is linked with 3rd
position. This forms phosphate diester bond. This bond links
carbon 5’ in one nucleoside with carbon 3’ in the next nucleoside.
3.Nirogenous bases: These are N2 containing organic compounds. They are of two types,
namely purines and pyrimidines.
Purines: Purines are two – ringed N2 compounds. They are of two types, namely adenine and
guanine.
Pyrimidines: These are single ringed N2 compounds. They are two types, namely thymine and
cytosine. Thymine and cytosine are the pyrimidine bases of DNA.
Structure of DNA (Watson and Crick model):
In 1933 Watson and Crik designed the structure of DNA. It is called the Watson and Crick model
of DNA. They were awarded with Nobel Prize in 1962 for this work. According to them DNA is
in the form of double helix. DNA is the deoxy ribonucleic acid. It is a nucleic acid. It is made up
of two chains. Each chain is the polynucleotide chain. Each polynucleotide is made up of many
small units called nucleotides. Each nucleotide is made up of three chemical components,
namely a phosphoric acid, a deoxiribose sugar and a nitrogen base. The nitrogen bases are
adenine, guanine, thymine and cytosine.
The nucleotides of DNA are named according to the type of nitrogen bases present. As there are
four types of nitrogen bases, DNA contain four types of nucleotides, namely
1.AMP- Adenosine monophosphate (adenylic acid)
2.GMP- Guanosine monophosphate (guanylic acid )
3. TMP- Thymidine monophosphate (thymodylic acid)
4. CMP- Cytidine monophosphate (cytidylic acid)
In each nucleotide, the deoxyribose sugar is attached to a phosphoric acid at one side and a
nitrogen base at the other side. The phosphoric acid is linked to the sugar. The nitrogen base
molecule is joined to the sugar by a glycosidic bond. This bond is formed between sugar and
nitrogen base.
Many nucleotides are linked together to form a polynucleotide chain. Two nucleotides are joined
by a phosphodiesterase bond. It is formed between sugar of one nucleotide and phosphate
component of another nucleotide.
The linking between purines and pyrimidines is brought about by hydrogen bonds. There are two
hydrogen bonds between A and T (A=T), and 3 hydrogen bonds between G and C (G= C). The
amount of adenine is equivalent to the amount of thymine and the amount of guanine is
equivalent to the amount of cytosine. The two chains of a DNA are complimentary to each other.
At one end of the polynucleotide chain, the 3rd
carbon atom of the sugar is free and it is not
linked to any nucleotide. This end is called 3 prime (3’) end. At the other end of the 5th
carbon of
the sugar is free and this end is called 5prime (5’) end.
DNA strand is antiparallel as they run in opposite direction. The DNA molecule is in the form of
a double helix. The two polynucleotide chains are coiled around each other to form a double
helix. The width (diameter) of DNA is 20A0. The DNA has two external grooves, namely major
groove and minor groove. The major groove is wider and deep. The minor groove is narrow. The
distance between two nucleotides is 3.4 A0.
Properties of DNA;
1. Size of the DNA molecule-The size of the DNA molecule varies from organism to organism.
It depends upon the size of the chromosome and the number of chromosomes found in each
living cell. The size basically depends upon the number of nucleotides present in each DNA
molecule. The size of DNA molecule ranges from 0.7 µm to 40,000mm (4cms).
2. Fragility of DNA molecule: The DNA molecule is highly fragile. Smaller DNA can be
isolated without any damage, but large sized DNA (above 2X 108 deltons) undergoes breakage
during their extraction.
3. Denaturation: Denaturation refers the separation of the two strands of a DNA. Denaturation
is brought about by high temperature, acid, or alkali. During denaturation there is breakdown of
hydrogen bonds between base pairs. Since G-C base pairs have 3 hydrogen bonds and A-T pairs
have 2 hydrogen bonds, G-C base pairs are more stable and it needs more temperature for
denaturation.
4.Renaturation: The denatured single stranded DNA can be made into double stranded DNA by
cooling or by neutralizing the medium. This process is called denaturation.
5.Effect of pH on DNA: The DNA is stable around the neutral pH in the solution. Further
increase in pH (alkali treatment) causes stand separation and finally denaturation occurs.
6.Stability : The DNA is a highly stable molecule. The stability is due to two forces.
a.( Hydrogen bonding between the bases. b.) Hydrophobic interactions between the bases.
Hydrogen bonds
7.Hyper chromic effect: DNA molecule absorbs light energy. This is a property of individual
bases. The intact DNA absorbs less light energy as its bases are packed into a double helix. A
denatured DNA molecule absorbs more light as its bases in single strands are exposed.
Functions of DNA: It plays an important role in all biosynthetic and heriditory functions of all
living organisms.
1.It acts as the carrier of genetic information from generation to generation.
2. DNA is a very stable macromolecule in almost all living organisms.
3.It controls all developmental processes of an organisms and all life activities.
4.DNA synthesizes RNAs.
5. DNA is the genetic code which is responsible for protein synthesis.
Nucleotides: Nucleotides are defined as phosphoric acid esters of nucleosides. A nucleotide is
made up of 3 components, namely a nitrogen base, a pentose sugar and a phosphoric acid.
The nucleotides are named according to purines and pyrimidines. AMP, GMP, TMP,CMP and
UMP. In addition many nucleotides occur freely in the tissues. They are ADP and ATP. On
hydrolysis nucleotide splits into phosphoric acid and a nucleoside. The nucleoside is made up of
a base and a pentose sugar. The hydrolysis of ATP yields ADP and energy.
Biological significance of nucleotides:
1. Nucleotides form the main components of nucleic acids.
2. Genetic material: Deoxyribonucleotides of DNA transmit hereditary characters from parents
to offspring.
3. Nucleotides functions as the source of high energy.Eg ATP, UTP, CTP, etc.
4. ATP is involved in oxidative phophorylation.
5. Certain nucleotides function as coenzymes. Eg UDPG, CoA, FMN, FAD.
6. Certain nucleotides function as vitamin B. Eg. FMN, FAD, NAD. Etc.
Nucleosides: Compounds that contain nitrogen bases linked to pentose sugars are called
nucleosides. There are two main types: ribonucleoside and deoxyribonucleoside. There are 5
types of nucleosides- adenosine, guanosine, thymidine, cytidine, uridine.
Types of DNA: DNA is classified into various types:
1.Double stranded DNA: It is also called as double helical DNA. In most of the organisms
except a few viruses, the DNA has a double stranded structure.
2.Single stranded DNA: Eg. Some viruses, E.coli, extra chromosomal satellite.
3. A-DNA: It is a double helical DNA having 11 residues per turn. It has a right handed helix. It
is formed by the dehydration of B.DNA.
A, B and Z DNA
4.B-DNA: This is the Watson and Crick double helix having 10 residues per turn. It is also right
handed. 5.Z- DNA: It is the left handed double helix having 12 residues per turn.
6. Circular DNA: It is circular in shape. It is found in bacteria, virus, mitochondria and
chloroplast. The circular DNA may be single stranded or double stranded. Single stranded
circular DNA is found in some viruses. Double stranded circular DNA is found in bacteria,
viruses, mitochondria, chloroplast, etc.
7. Relaxed DNA: Circular DNA without any helical coiling is called relaxed DNA.
8. Supercoiled DNA: It is supercoiled DNA. It can produce negative super coiling and positive
supercoiling. The degree of supercoiling is controlled by topoisomerases and gyrases.
9.Palindromic DNA: A double helix is formed by two paired strands of nucleotides that run in
opposite directions in the 5I- to 3I sense and the nucleotides always pair in the same way A-T for
DNA , with Uracil (U) for RNA, Cytosine ( C ), a nucleotide said to be palindrome if it is equal
to its reverse complement. For example, the DNA sequence ACCTAGGT is palindromic
sequence because its compliment is TGGAGGT. The sequence of nucleotide goes in one
direction and in another direction in the second strand.
10. Repetitive DNA or satellite DNA: When very short sequences of base pairs are repeated
many times in DNA, the DNA is called repetitive DNA or satellite DNA. All eukaryotes, except
yeast, contain repetitive DNA. Repetitive DNA is absent in prokaryotes. The repetitive DNA can
replicate but cannot transcribe mRNA for protein synthesis. Repetitive DNA is therefore inert.
Ribonucleic acid (RNA): It is a nucleic acid containing ribose sugar. It is found in large amount
in the cytoplasm and at a lesser amount in the nucleus. In the cytoplasm it is found mainly in the
ribosomes and in the nucleus it is mainly found in the nucleolus. RNA is formed of a single
strand. It consists of several units called ribo-nucleotides. Hence each RNA molecule is formed
of several nucleotides. Each nucleotide is formed of different molecules, namely phosphate,
ribose sugar and nitrogen base. The nitrogen bases are purines and pyrimidines. The purine bases
present in the RNA are adenine and guanine. The pyrimidines present in the RNA are cytosine
and uracil. The RNA molecule is normally single stranded, sometimes the stand may be folded
back upon itself and this double strand may be coiled to form a helical structure like that of
DNA. In RNA purines and pyrimidines are not present in equal amount.
RNA structure
There are three types of RNA. They are mRNA, tRNA and rRNA
mRNA: This type of RNA carries genetic information for protein synthesis from the DNA to the
cytoplasm. The mRNA forms about 3 to 5% of the total cellular RNA. The mRNA is synthesized
as a complimentary strand upon the chromosomal DNA. The genetic message from DNA is
transcribed in this hybrid mRNA. The mRNA carries the message in the form of triplet codes.
The hybrid mRNA inside the nucleus is called heterogenous nuclear RNA (hnRNA). It is
processed in the nucleus and enters the cytoplasm through nuclear membrane. In the cytoplasm
mRNA are deposited on some ribosomes. In the ribosomes mRNA acts as a template for protein
synthesis.
The life span of mRNA in bacteria is about 2 min. In eukaryotes it lives for few hours to a few
days.In the animal eggs and a plant seed, the mRNA is stabilized for months or years. Protein
synthesis must be carried within this life span.
Types of mRNA: There are two types of mRNA
a)Monocistronic mRNA: It is formed from a single cistron (functional gene). It contains the
genetic information to translate only a single protein chain (polypeptide). The eukaryotic mRNAs
are monocistronic.
b) Polycistronic mRNA: It is formed from several cistrons. The polycistronic mRNA carries
several open reading frames (ORFs), each of which is translated into a polypeptide. Hence
polycistronic mRNA translate several polypeptide chains. The prokaryotic mRNA are
polycistronic.
Structure of mRNA: It is a single stranded polynucleotide chain. Each nucleotide is made up of
many nucleotides. Each nucleotide contains a phosphoric acid, a ribose sugar and a nitrogenous
base. The nitrogenous base may be adenine or guanine or cytosine or uracil.Among RNAs,
mRNA is the longest one. Most of the mRNAs contain 900 to 15000 nucleotides. One end of the
mRNA is called 5’ end and other end is called 3’ end. At the 5’ end a cap is found in most
eukaryotes and animal viruses. The cap is formed by the condensation of the guanylate residue.
The cap helps the mRNA to bind with ribosomes.
mRNA
The cap is followed by a non-coding region1 (NC1). It does not contain code (message) for
protein and hence it cannot translate protein. It is formed by 10 to 100 nucleotides and is rich in
A and U residues. The non-coding region is followed by the initiation codon. It is made up of
AUG. The initiation codon is followed by the codon region which contains code for protein. It
has an average 1,500 nucleotides. The codon region is followed by a termination codon. It
completes the translation. It is made up of UAA or UAG or UGA in eukaryotes. The termination
codon is followed by non-coding region 2(NC2). It has a nucleotide sequence of AAUAAA. At
the 3’ end of mRNA, there is a polyadenylate sequence (poly A). It consists of 200 to 250
adenylate nucleotides (AAAAA….). But as the age increases, the poly A shortens.
The mRNA is synthesized from a DNA strand through the action of an enzyme called RNA
polymerase. The synthesis of mRNA is called transcription. mRNA carries the genetic
information from the DNA. The code decides the type of protein to be synthesized.
The mRNA carries genetic information from DNA. The genetic information carried by the
mRNA is called genetic code. The genetic code is the sequence of nitrogen bases in mRNA. The
genetic code is formed of several codons. Each codon is a sequence of three nitrogen bases
which codes for one aminoacid. As each codon is formed of threenucleotides, it is called a triplet
code. Each mRNA contains the codons for one polypeptide chain. It the mRNA contains 900
nucleotides the polypeptide chain synthesized by this mRNA will contain 300 aminoacids.
Transfer RNA (tRNA): It is a ribonucleic acid which transfers the activated aminoacids to the
to the ribosomes to synthesize proteins. It is so small that it remains in the supernatant during
centrifugation. Hence it is also called soluble RNA or supernatant RNA. It serves as an adaptor
molecule to attach amino acids. Hence tRNA is also called adaptor RNA. It constitutes 10 to
15% of the total weight of RNA of the cell. It has a molecular weight of 25000 to 30000 and a
sedimentation co-efficient of 3.8S.
Structure of tRNA: The tRNA is made up of 73 to 95 nucleotide units called ribonucleotides.
Each nucleotides unit is made up of three components, namely a phosphate, a ribose sugar and a
nitrogenous base such as adenine, guanine, cytosine or uracil. The tRNA is in the form of single
polynucleotide chain having 3’ and 5’ ends. The polynucleotide chain of tRNA is folded on itself
and attains the shape of clover leaf. The 3’ and 5’ ends of tRNA lie side by side as a result of
folding. The 3’ end always ends in CCA base sequence. This is the site for the attachment of
activated amino acid. The 5’ end terminates in G or C. The t RNA has 5 arms.
a.Amino acid acceptor arm, b) D-arm, c) Anticodon arm, d)Variable arm, e) T ΨC arm. Each
arm is made up of a stem and a loop. But the acceptor arm has no loop; the variable arm has no
stem. In the stem, the bases pair with each other (A-U and G-C). There is no base pairing in the
loops.
a) Amino acid acceptor arm: In the amino acid acceptor arm, the stem does not end with loop.
The acceptor has 3’ end of the nucleotide chain. The terminus of the acceptor site has a constant
CCA base sequence. To this base amino acids are attached to form aminoacyl tRNA. The 5’ end
of the arm comes near the 3’ end due to folding. Its terminus is either G or C.
b) D.arm: The D arm has 3 to 4 bases in the stem and 7 to 11 unpaired bases in the loop. The
loop is called dihydrouridine loop (DHU) or D-loop.
c)Anticodon arm: In the anticodon arm, the stem has 5 paired bases and the loop has 7 unpaired
bases. The loop is called anticodon loop. Three of the 7 unpaired bases in the loop (anticodon)
determine the pairing of tRNA with the specific codon of mRNA.
d) Variable arm: In variable arm the stem may or may not be formed. The variable arm or mini
arm has a loop with 4-5 bases.
e) T ΨC arm: The T ΨC arm contains a constant T ΨC sequence. Its loop has ribosome
recognition site.
The tRNA molecules are named according to the amino acid to which it gets attached. For
example, tRNA carrying alanine can be called tRNAal. The tRNA molecules are synthesized, at
particular regions of DNA by a process called transcription. About 40 to 80 genes or cistrons are
involved in tRNA transcription.. The hybrid tRNA has base sequences complimentary to the
mother DNA in the beginning. But, after the completion of transcription, the nitrogenous bases
are altered at certain points in the nucleotide chain.
Functions of tRNA: tRNA picks up a specific activated amino acid from the amino acid pool in
the cytoplasm (aminoacyltRNA). The amino acid is then transferred to the ribosome in the
cytoplasm where the proteins are synthesized. The attachment with ribosome depends upon the
codes in the mRNA and anticodons in the tRNA. Finally it transmits its amino acid to the new
polypeptide chain.
Ribosomal RNA (rRNA): It is a ribonucleic acid present in the ribosomes and hence it is called
ribosomal RNA. It is also called insoluble RNA. It constitutes about 80% of the cellular RNA. It
is formed by a single strand. It is a polynucleotide chain. Each strand is formed of many
nucleotide units. Each nucleotide is formed of three components- a phosphare, a ribose sugar and
a nitrogen base. The purine bases present in rRNA are adenine and guanine and the pyrimidine
bases are cytosine and uracil. Each strand has a 5’ end and a 3’ end. In some regions, the single
strand is twisted upon itself to form a double helix. In the helical regions most of the base pairs
are complimentary. They are joined by hydrogen bonds. In the unfolded single stranded regions,
the base pairs are not complimentary. In prokaryotes, the rRNA has 3 types: 23S, 5S, and 16S. In
the mammals, 4 types of rRNA have been found: 28S, 5.8 S, 5S and 18S. The unit S strand for
Svedberg, which is a measure of the sedimentation rate. After rRNA molecules are produced in
the nucleus, they are transported to the cytoplasma, where they combine with specific proteins to
form a ribosome. In prokaryotes, the size of a ribosome is 70S, consisting of two subunits: 50S
and 30S. The size of a mammalian ribosome is 80S, comprising a 60S and a 40S subunit.
Proteins in the larger subunit are designated as L1, L2, L3, etc.(L=large). In the smaller subunit,
proteins are denoted by S1, S2, S3, etc.
Composition of rRNA
During protein synthesis, the ribosome binds to mRNA and tRNA as above. Only the tRNA
containing the anticodon which matches mRNA codon may join the complex.
Functions of rRNA; It forms the main bulk of the cytoplasmic RNA. Its function is not clearly
known. However it is believed that rRNA plays the major role in protein synthesis.
c) DNA replication: General, bacterial and eukaryotic DNA replication. Replication is the duplication of DNA. By replication DNA produces exact copies of its own
structure. Replication occurs inside the chromosomes. It occurs during interphase. The parent
DNA strands function as templates for the synthesis of new DNA strands. New DNA is produced
by semi conservative process. Of the two strands produced, one strand is the parental strand and
the second strand is newly synthesized.
Replication starts at a specific point called origins. At this origin, the two strands are separated.
This separation is brought about by the enzyme helicase. At the point where the two strands are
separated, a replication fork is formed. The fork appears in the form of Y. The duplication of
DNA is brought about by the movement of the replication fork. Single stranded DNA binding
proteins or SSB, binds to single-stranded regions of DNA to prevent premature annealing, to
prevent the single-stranded DNA from being digested by nucleases and allow other enzymes to
function effectively upon it.
Leading strand- In one DNA strand, the daughter strand is synthesized as a continuous strand.
This strand is called leading strand because it is synthesized first. The leading strand is
synthesized continuously in the 5’ 3’ direction by DNA polymerase.
DNA replication Bidirectional replication
On the leading strand, a DNA polymerase “reads” the DNA and adds nucleotides to it
continuously. This polymerase is DNA polymerase III in prokaryotes. In human cells the leading
and lagging strands are synthesized by Pol α and Pol δ within the nucleus and Pol γ in the
mitochondria.
RNA primer- A primer is a strand of nucleic acid (10-12 nucleotides) that serves as a starting
point for DNA synthesis. They are required for DNA replication because the enzymes that
catalyze this process, DNA polymerases can only add new nucleotides to an existing strand.
The polymerase starts replication at the 3’ end of the primer and copies the opposite end. In most
cases of natural DNA replication, the primer for DNA synthesis and replication is a short strand
of RNA. The RNA primer is synthesized by DNA dependent RNA polymerase (primase). The
RNA primer is degraded at the end of DNA replication.
Lagging strand- In the second DNA strand, the daughter strand begins slightly later. Hence this
daughter strand is called lagging strand. The lagging strand is synthesized in short fragments
called Okazaki fragments. The replication is semi conservative discontinuous because the DNA
is synthesized in short segments. The enzyme DNA ligase joins the Okazaki fragments into a
long polynucleotide chain. DNA ligases catalyze formation of a phosphodiester bond between
the 5’ phosphate of one strand of DNA and the 3’ hydroxyl of another.
Replication may occur in one direction from the point of origin or in both directions. When
replication occurs in only one direction, it is called unidirectional replication. When replication
occurs in both directions, it is called bidirectional replication.
Bacterial (prokaryotic) DNA replication- DNA replication in E.coli begins at a single, defined
DNA sequence of 245 base pairs called oriC . A protein called DnaA gets concentrated near oriC
and get complexes with ATP. This complex binds with specific 9-bp repeats at oriC. These
distors the DNA, leading to the opening of adjacent 13- bp repeats in the DNA. The opened
DNA strand allows protein complex to enter the DNA bubble. The protein complex consists of
DNA helicase (Dna-B) and a DNA helicase loader (Dna-C). DNA helicase loader open the
DNA helicase protein rings and place the rings around the single stranded DNA. The loaders are
then released. The helicases use energy from ATP hydrolysis to unwind the DNA helix at each
of the two replication forks. Each DNA helicase recruits an enzyme called DNA primase,(DNA
dependent RNA polymerase) which synthesizes RNA primer on the DNA template. The RNA-
primer contains 10-12 nucleotides.
The main replication enzyme in E.coli is called DNA polymerase III. This enzyme synthesizes
the new DNA 5’ to 3’ direction. This is leading strand. In contrast, the other new strand called,
called the lagging strand is built in fragments called, Okazaki fragments. The template strands
are antiparallel, with their 3’ and 5’ ends oriented in opposite directions. Many proteins
participate in DNA replication. Single- strand DNA binding proteins quickly coat the exposed
single stranded DNA. These proteins protect the DNA strand against nucleases. DNA replication
continues as the DNA polymerase on the lagging strand meets the 5’ end of the next primer.
After the DNA helicase has moved approximately 1000 bases, a second RNA primer is
synthesized at the fork. The cycle continues for the length of the template strands. The lagging
strand consists of Okazaki fragments with a segment of RNA at one end. The RNA is cleaved by
an enzyme called RNase H. Another enzyme DNA polymerase I fills the large gap between
Okazaki fragments. Finally DNA ligases catalyze formation of a phosphodiester bond between
the 5’ phosphate of one strand of DNA and the 3’ hydroxyl of another. During replication, DNA
can become super coiled. DNA gyrase (topoisomerase II) prevents this super coiling.
Topoisomerase IV separates the newly formed daughter circular DNA strand from the parent.
Eukaryotic DNA replication: DNA replication in eukaryotes is much complicated in
prokaryotes, although there are many similar aspects. Eukaryotic cells only initiate DNA
replication at a specific point in the cell cycle- the beginning of the S phase. However, pre-
initiation occurs in the G1 phase.
d) The cell cycle: Restriction point, cell cycle regulators and modifiers.
Cell cycle: (cell-division cycle) It is the series of events that takes place in a cell leading to its
division and duplication (replication). In prokaryotic cells, the cell cycle occurs via a process
termed binary fission. In eukaryotic cells, the cell cycle can be divided in two periods- a)
interphase, b) mitosis
a) Interphase- During this phase the cell grows, accumulating nutrients needed for mitosis and
duplicating its DNA.
b) Mitosis (M)-phase- During which the cell splits itself into two distinct cells
The duration of the cell cycle varies from hours to years. For example, the cell cycle of
Paramecium aurelia has duration of 6h. A typical human cell has duration of 90h.
a)Interphase: It is the longest phase. In a typical human cell, out of the 90h, interphase lasts for
89h.
Resting (Go phase): In interphase the cell prepares itself to cycle. The term post-mitotic is
sometimes used to refer both quiescent and senescent cells. Non-proliferative cells in eukaryotes
generally enter the quiescent Go state from G1 and may remain quiescent for longer period of
time or indefinitely (e.g.cardiac cells and neurons). In multicellular eukaryotes, cells enter the Go
phase from the G1 phase and stop dividing. Some cells enter the Go phase semi-permanently, e.g.
some liver and kidney cells.
Characters of interphase: It is the resting phase of the cell. Resting refers to the rest from
division. But, the cells in the interphase are metabolically active. The metabolic activities are
high in this phase. The cell grows during phase. During this phase mRNA and rRNA are
synthesized. Throughout the interphase the chromosomes are extended and are not visible in the
light microscope. The chromosomes duplicates into two chromatids. The centrioles duplicates
into two. Thus two centrioles are formed. The centrospheres of centrioles, microtubules arise.
These microtubules form asters.
Stages of interphase: Interphase consists of 3 sub-stages. They are G1 phase, S phase and G2
phase.
G1 Phase: G stands for gap. It is the first phase within the interphase, from the end of the
previous M phase. It is also called the growth phase. This phase is the gap period between a
mitotic phase and the S phase of the cycle. This period starts immediately after division. The
daughter cells grow and increase in size during this phase. It is a longer phase. It lasts for even
years. The nerve cells remain permanently in G1 phase. Generally, this stage lasts for 25 to 50%
of the total interphase. During this phase 20amino acids are formed, from which millions of
proteins and enzymes are formed, which are required in S phase. During this phase mRNA,
rRNA and tRNAs are formed. During this phase new cell organelles are formed.
G1 phase consists of four sub-phases: Competence (g1a), entry (g1b), progression (g1c) and
assembly (g1d). These sub-phases may be affected by limiting growth factors, nutrient supply,
temperature, and additional inhibiting factors. A rapidly dividing human cell which divides every
24h spends 9h in G1 phase. The DNA in a G1 diploid eukaryotic cell is 2n, meaning there are two
sets of chromosomes present in the cell. The genetic material exists as chromatin.
There is a restriction point present at the end of G1 phase. Signals from extracellular growth
factors are transducer in a typical manner. Growth factors bind to receptors on the surface.
Accumulation of cyclin D’s is essential. Cyclins are a family of proteins that control the
progression of cells through the cell cycle by activating cyclin-dependent kinases (Cdk)
enzymes. Cyclin D acts as a mitogenic signal sensor.
S-phase: S stands for synthesis: During this phase DNA synthesis occurs. The DNA molecule
duplicates. All the chromosomes have been replicated. This period lasts for 35 to 40% of
interphase. During this phase, synthesis is completed as quickly as possible due to the exposed
base pairs may be destroyed by the external proteins (drugs) or any mutagens (such as nicotine).
Cyclins, when bound with the dependent kinases such as Cdk1 proteins form the maturation-
promoting factor (MPF). MPFs activate other proteins through phosphorylation. These
phosphorylated proteins, in turn, are responsible for specific events during cell division such as
microtubule formation and chromatin remodeling.
DNA damage checkpoints: These sense DNA damage both before the cell enters S phase (a G1
checkpoint) as well as after S phase (a G2 checkpoint). Damage to DNA before the cell enters S
phase inhibits the action of Cdk2 thus stopping the progression of the cell cycle until the damage
can be repaired. If the damage is so severe, that it cannot be repaired, then the cell destructs by
apoptosis. Damage (UV radiation, oxidative stress, etc) to DNA after S phase (the G2
checkpoint), inhibits the action of Cdk1 thus preventing the cell from proceeding from G2 to
mitosis. In the S phase if DNA replication stops at any point on the DNA, the progress through
the cell cycle is halted until the problem is solved.
G2 phase (pre-mitotic phase) The G2 phase is the gap period between S-phase and mitotic (M)
phase of a cell cycle. It is the second growth phase. It is a period of rapid cell growth and protein
synthesis which the cell readies itself for mitosis. The nucleus increases in volume. Metabolic
activities essential for cell division, occur during this phase. mRNA, tRNA and rRNA synthesis
also occur. It is not a necessary part of the cell cycle. The G2 phase is followed by mitotic phase.
b)M-phase (Mitotic phase): This is the division phase. During this phase the cell divides. This
phase has a short duration. A typical human cell cycle has duration of 90h. Of these the M phase
has duration of 45 to 60min. This phase has two sub-phases called karyokinesis and cytokinesis.
Karyokinesis refers to the cell division of nucleus into two daughter nuclei. It has 4 sub-stages,
namely prophase, metaphase, anaphase and telophase, Cytokinesis refers to the cell division of
the cytoplasm resulting in two daughter cells.
Cell cycle checkpoints (Restriction points): These are the cell cycle control mechanisms in
eukaryotic cells. These checkpoints verify whether the processes at each phase of cell cycle have
been accurately completed before progression into the next phase. There are three main
checkpoints that control the cell cycle in eukaryotic cells. They are
1.G1 checkpoint (G1restriction point)
2.G2 checkpoint
3.Metaphase checkpoint.
1.G1 checkpoint (G1restriction point): This checkpoint is present at the end of the G1 phase
and just before of the S phase of the cell cycle. This checkpoint helps in taking the decision of
whether the cell should divide, delay division, or enter a resting stage (Go phase). If there are
unfavorable conditions for the cell division, then this restriction point restrict the progression to
the next phase by passing the cell to Go phase for an extended period of time.
This restriction point is controlled mainly controlled by the action of the CKI-p16 (CDK
inhibitor p16). The inhibited CDK not bind with cyclin D1, hence there is no cell progression.
The E2F group of genes (transcription factors) that are present in eukaryotic cells, they control
the progression of cell cycle. Transcription activators such as E2F1, E2F2, and E2F3a promote
the cell cycle, while repressors like E2F3b, E2F4, E2F5, E2F6, E2F7, and E2F8 inhibit the cell
cycle. This restriction point is overcome by the increased expression of cyclin D, which then
interact with CDK4/6 phosphorylate the tumour suppressor retinoblastoma (Rb), which relieves
the inhibition of the transcription factor E2F. E2F is then able to cause expression of cyclin E,
which then interacts with CDK2 to allow for G1-S phase transition. This brings the cell to the end
of the first checkpoint.(unphosphorylated Rb inhibits the E2F).
2.G2 checkpoint: This restriction point is located at the end of the G2 phase. This checks the
number of factors which are essential for the cell division. Maturation-promoting factor or
mitosis promoting factor or M-phase promoting factor- (MPF) is a protein composed of cyclin-B
and CDK-1. This protein promotes the G2 phase into the entrance of M-phase. MPF is activated
at the end of G2 by a phosphatase (Chk) which removes an inhibitory phosphate group added
earlier.
The main functions of MPF in this restriction point are
a.Triggers the formation of mitotic spindle.
b.Promotes chromosome condensation.
c.Causes nuclear envelop breakdown.
If there are any damages are noticed in this restriction point, then phasphatase not activate the
MPF, resulting in the arrest of cell cycle in G2 phase till the repair of the damaged DNA. This
prevents the transfer of defected DNA into the daughter cells.
3.Metaphase checkpoint: This occurs at metaphase. Anaphase-promoting complex (APC)
regulates this checkpoint. This is also called spindle checkpoint. This checks whether all
chromosomes are properly attached to the spindle or not. This also governs the alignment of the
chromosomes and integrity of the spindles. If there are mistakes then it delays the cell in entering
into anaphase from metaphase.
Cell cycle regulators: The cell cycle is regulated by cycles, cyclin-dependent kinsases (CDKs),
and cyclin-dependent kinase inhibitors (CDKIs).
1.Cyclins: Their concentration varies during the cell cycle. Cyclins are the family of proteins
which regulates the cell cycle. There are several types of cyclins that are active in different parts
of the cell cycle and causes phosphorylization of CDK. There are also several”orphan” cyclins
for which no CDK partner has been identified. For example, cyclin F is an orphan cyclin that is
essential for G2/M transition. There are two main groups of cyclins:
a.G1/S cyclins: Examples- Cyclin A,D and E. These cyclins are essential for the control of the
cell cycle at the G1/S transition. CyclinA/CDK 2- active in S phase. Cyclin A binds to S phase
CDK 2 and is required to progress through the S Phase. Cyclin A/CDK 2 is inhibited by the
complex p21CIP.
The un-phophorylated form of Rb binds with E2F family of transcription factors which controls
expression of several genes involved in cell cycle progression (example-cyclin-E). Rb acts as a
repression, so in complex with E2F it prevents the expression of E2F genes, and this inhibits the
cell progression from G1 to S phase. The binding of cyclin D/CDK4 and cyclin D/CDK 6 lead to
partial phosphorylation of Rb, by reducing its binding to E2F. The E2F gene activates the
expression of cyclin E bind with CDK 2 and causes complete phosphorylation of Rb. This
progresses the cell cycle from G1 to S phase.
a.G2/M cyclins: Example; cyclin B/ CDK1-. These are essential for the control of the cell cycle
at the G2/M transition. G2/M cyclins accumulate steadily during G2 and are abruptly destroyed as
cells exit from mitosis (at the end of the M-Phase). Cyclin B/ CDK1- regulates progression from
G2 to M Phase. Cyclin B is a mitotic cycin. The amount of cyclin B (which binds to CDK1) and
the activity of cyclin B-CDK complex rise through the cell cycle until mitosis. The cyclinB-
CDK1 complex is called maturation promoting factor (MPF). There are two types of cyclin B.