1
(CHROMOSOMES AND DNA)
History
In 1848, Hofmeister observed filament or thread like structures in the nuclei of pollen mother cells of T radescantia.
The German embryologist Walther Flemming observed these threads in 1882, when he was examining the rapidly dividing
cells of salamander larvae. In 1888 Waldyer name them chromosomes which means coloured body (Greek, Chromoe means
color and Soma means body) because these were darkly stained. Chromosomes are found in the cells of eukaryotes. Most
nuclear DNA is attached to proteins, forming very long fibers called chromatin. During cell division chromatin condenses
into chromosomes. The number of chromosome varies from species to species. The number may vary from two e.g. Mucor
to more than 500 pairs in some other organisms.
Types of chromosomes
A chromosome is marked by a constriction known as primary constriction. The primary constriction divides the
chromosme into two arms. In the region of primary constriction lies the centromere. It is associated with granule like
structure made of proein the kinetochore, to which spindle fibres attach during cell division. A chromosme mey have one
more constriction. It is called secondary constriction. The tip of the chromosomes are rounded and sealed and are called
telomeres. The terminal part of the chromosme beyond secondary constriction is called satellite. Based on the location of
primary constriction and kinetochore, the chromosomes are described as:
Metacentric: These are V-shaped chromosomes, have centromere in the middle so that the two arms of the
chromosomes are almost equal.
Submetacentric: These are L-shaped chromosomes with centromere slightly away from the midpoint so that the two orms
of the chromosme are almost unequal.
Acrocentric: These are rod shaped chromosomes with terminal centromere, so that the possess only one arm.
Telocentric: These are also rod-shaped but with subterminal centromere have one arm very long and the other arm is
very short.
Karyotype
When chromosomes are arranged by pairs according to their size, shape and general appearance in mitotic
metaphase it is called karyotype. It is the total chromosomal complement of a cell. Study of karyotype is used for the
identification of chromosomal number. The figure number 20.3 outlines the steps in one method of preparation of a
karyotype from a blood sample. Karyotyping is used to screen the abnormal number of chromosomes and for defective ones.
Chemical Composition of Chromosomes
Chemically chromosomes are composed of nearly 40% DNA and 60% protein. A significant amount of RNA is
also associated with chromosomes. A typical human chromomsome contains about 140 million nucleotide of DNA.
Ultra Structure of Chromosmes
Eukaryotic chromosomes contain an enormous amount of DNA relative to their size. Each one contains a single
DNA molecule that is thousands of times longer than the 5 mm diameter of a typical nucleus. All this DNA can fit into the
nucleus because of an elaborate, multilevel system of coiling and folding of the DNA in each chromosome.
How is the coiling of long DNA fiber achieved?
Nucleosomes: DNA is associated with basic protein molecules called histones. DNA and histone become organized into
nucleoprotein fibers. When stretched out a nucleoprotein looks like a beaded chain under the electron microscope. Each
“bead” in a chain is a nucleosome. It is 10 nm in diameter. Each nucleosome is an aggregate of histone molecules
(octamere), of two type (Fig 20.4). The nucleosome is repeated after every 200 nucleotides of the DNA.
Spacer or linker DNA: The nucleosome “beads” in a chromatin appear to be held together by segments of DNA called
linker DNA or spacer DNA, that stretch from one nucleosome to the next.
Super coils: Further coiling of the DNA occurs, when the string of nucleosome warps into higher order coils called super
coil, (200 nm diameter)
Chromatin:
The super coil network is called chromatin. Chromatin forms super coils within the chromosomes. Highly
condensed portions of chromatin is called heterochromatin. Some of these portions remain permanently condensed. The part
of the chromosme which is condensed only during cell division is called euchromatin.
Chromosome theory of inheritance:
Mendel competed his results in 1866, American genetist Karl Correns rediscovered Mendel’s work in 1900 and
suggested the central role for chromosme in heredity. In 1902 Walter S. Sutton noticed the similarities between the behavior
of chromosomes during the formation of gametes and fertilization of Mendel’s hereditary factor. W.S Sutton of U.S.A and
Theodor Boveri of Germany suggested independently that the chromosomes were the carrier of Mendel’s hereditary factors,
the chromosme throry of inheritance.
The chromosome theory states that genes are located on chromosomes and that the behaviour of chromosomes
during meiosis and fertilization accounts for inheritance patterns. The chromosomes undergo segregation and independent
assortment during meiosis and thus, account for Mendel’s principles.
We can see the chromosomal basis of Mendel’s principles by following the fate of two genes during meiosis and
fertilization in plants. In the figure No. 20.6 pea the genes for seed shape (alleles R and r) and seed color (alleles Y and y)
are shown as black bars on different chromosomes. We start with the F 1 generation, in which all individuals have the RrYy
genotype, to simplify the diagram, we show only two of the seven pairs of pea chromosomes and three of the stages of
meiosis: metaphase I, anaphase I, and metaphaseII.
To see the chromosomal basis of the principle of segregation, let’s follow just the pair of long chromosomes, the
ones carrying R and r, taking either the left or the right branch from the F 1 cell. Whichever arrangement the chromosomes
assume at metaphase I, the two alleles segregate as the homologous chromosomes, separate in anaphase I. and at the end of
meiosis II, a single long chromosme ends up in each of the gametes. Random fertilization then leads to F 2 offspring with the 3:1
(12 round to 4 wrinkled) ratio of phenotypes that Mendel observed.
(CHROMOSOMES AND DNA)
2
To see the chromomal basis of the principle of independent assortment, follow both the ,ong and the short
chromosomes from metaphase I in the F1 generation. Two alternative, equally likely arrangements of tetrad can occur at this
stage of meiosis. The nonhomologous chromosomes and the genes they carry assort independently, forming gametes of four
genotypes. Random fertilization leads to the 9:3:3:1 phenotypic ratio in the F 2 generation.
Table 20.2 Similarities between the Behavior of Genes
and Chromosomes
Behaviour of genes Behaviour of
Chromosomes
1) The genes that control 1) Diploid cells contain the
the particular characteristic pair of chromosomes
occur in pairs. (homologous chromosomes)
2) Pairs of the genes 2) Each gamete cell has
separate during the gamete only one out of a
formation so that the homologous pair of
gametes have only one of chromosome i.e. gametes
the paired again. are haploid.
3) When the genets unite 3) When the gametes unite
the gene pairs are formed during the fertilization,
again. chromosomes again occur in
pair. The zygotes are
diploid.
4) Genes are transmitted 4) Individual chromosomes
from one generation to the are transmitted from
next and remain as generation to generation as
unchanged units. unchanged structure.
5) There is independent 5) There is independent
assortment of genes i.e. assortment of chromosomes
possible gene combination i.e. possible chromosomes
can be calculated. combination can be
calculated.
Search for the Hereditary Material
In 1928 Frederick Griffith performed experiments on the bacteri namely Streptococcus pneumoniae (the known as
Pneumococcus) that cause the pneumonia disease. The bacteria can exist in two different forms. Two types of bacteria were
isolated from diseased mouse:
(1) Smooth strain of bacteria (S)
(2) Rough strain bacteris (R)
Smooth strain of bacteria were virulent (cause disease) and are covered with a polysaccharide capsule. Rough strain
of bacteria have no capsule and are called non-virulent because they were not able to cause pneumonia.
Experiments of Griffith
(i) Living R strain of bacteria were injected into a living mouse.
Result: the mouse remained alive.
(ii) Living S strain of bacteria were injected into the mouse.
Result: The mouse died.
(iii) Heat killed S strain of bacteria were injected into the mouse.
Result: Mouse remained alive.
(iv) A small number of living non virulent R strain and a large number of heat killed S strain of bacteria were injected
into the mouse.
Result: Mouse died of pneumonia.
The blood of the mouse was examined. A large number of virulent S strain of bacteria were round. The R type were
transformed into S type virulent bacteria. Something in dead cells had converted the living R cells material from one cell to
another and can alter the genetic make up of the recipient cell.
Work of T. Avery and His co workers on Transforming Principle
In 1944 Oswald Avery and his co-workers Colin Macleod and maclyn McCarty worked on transforming principle.
Avery with his co workers isolat4ed pure DNA from the Pneumococcus (Streptococcus pneumonias). When DNA in the
sample was destroyed, the transforming activityu was lost. It shows that DNA is the genetic material in the cell.
Alfred Hershey and Martha Chase
In 1952 Alfred Hershey and Martha Chase proved that the DNA is the heredity material performing experiment on
the phage virus T2. The virus that reproduce inside the body of bacteria are called phage virus or bacteria phage.
In the phage viruse phosphorus is present only in DNA and sulphur is present only in the protein. The protein and
DNA could therefore differentially tagged with radioactive elements. Two groups of bacteria E. coli were taken. AGroup A
was grown in medium S. The radioactive elements were absorbed and became a part of bacteria. Viruses were grown on
these bacteria. As a result one group of virus were labeled with 32P and other group with 35S.
These viruses were grown on ordinary bacteria having no radioactivity. They reproduced inside the bacteria. The
offspring came out bursting the wall of the bacteria. The offspring were tested of radioactivity. 32P was present in Group A
bacteria and 35S was absent in group B bacteria. It proved that only DNA entered the bacteria and reproduced. So this
experiment proved that DNA is the heredity material.
Chemical Composition of DNA
(CHROMOSOMES AND DNA)
3
Friederick Mieschr: He was a german biochemist. He isolated a substance in 1869. He name the substance as nuclein,
because it was located in the nucleus of the fish sperm cells. Nuclein was later on called nucleic acid, as it was acidic.
Work of Levene: In 1920 the basic structure of nucleic acids was determined by the biochemist P.A. Levene. He found that
DNA contains three main components (i) Phosphate group (ii) Five carbon sugars (iii) nitrogen containing bases called
purine i.e. Adenine, Guanine, and pyrimidine i.e. Thymine, Cytosine. RNA contains Uracil instead of thymine. Levene
concluded that DNA and RNA are made of repeating units called nucleotide.
Erwin Chrgaff: Tevealed that amount of Adenine was exactly matched with the Thymine and Guanine was equal in
amounts to Cytosine in DNA i.e there is always equal proportion of purine and pyrimidine.
Mauric Wilkins and Rosalind Franklin: Mauric Wilkins prepared highly oriented DNA fibres for study, using X-ray
crystallography. In this technique an X-ray bean is passed through a crystal of the substance being studied. Part of the X-ray
beam is scattered (diffracted) as it passes through the crystal. The way in with it scatters depends upon the structure of the
molecule. A photographic plate on the other side of the crystal records a paterns of spots representing the intensity of the
emergent X-rays. This pattern reveals information about the locations of various atoms in the crystals, which in turn, can be
used to determine the three dimensional shapes of molecules.
Wikins prepared DNA fibers. One of his colleague was Rosalind Franklin. She produced X-ray crystallographic
photograph of DNA. Watson saw the photographs. The photograph clearly revealed the basic shape of DNA to be helix. On
the basis of Waston’s later recollection of the photograph, he and Crick deduced that the helix had a uniform diameter of
2nm. The diameter of the helix suggested that it was made up of two polynucleotide strands. The presence of two strands
accounts for the now familiar double helix.
Structure of Nucleotide
A base is a nitrogen containing ring. It is called base because of unshared pair of electrons on the nitrogen atoms,
which can thus, acquire a proton. The base are purine and pyrimidine. Purine are Adenine and Guanine, which are double
ring structure. Pyrimidine are cytosine,Thymine and uricil which are single ring structure. When a base is linked with sugar
it is called nucleoside when a phosphate is linked with a nucleoside it is called nucleotide.
Nitrogen base is attached to carbon number 1 of a pentose sugar and a phosphate group is attached to carbon
number 5' of the sugar. In addition a free hydroxyl (OH) group is attached to 3’ carbon atom. The nucleotides are named
after the name of the base attached to it e.g. Adenine nucleotide. (Adenine deoxy-ribose phosphate). Bases are represented
by their initial letters A,G,T,C and U.
Polynucleotide
Each nucleoside in the polymer of DNA is linked to neighboring nucleosides by phosphate groups. These
phosphates connect the 3’ carbon of one sugar with the 5’ carbon of the adjacent nucleoside sugar. The reaction between 5’
phosphate group of one nucleotide and 3’ hydroxyl group of another is a dehydration synthesis, eliminating a water molecule
and forming a covalent bond. The linkage is called a phosphodiester bond because the phosphate group is now linked to the
two sugars by means of a pair of ester (P-O-C) bonds. Nucleotides can join together forming a long a long polyptide chain.
The structure of a RNA strand is similar except the ribose replaces the deoxyribose. Linear strands of DNA or RNA
no matter, how hong, will almost have a free 5’ phosphate group at one end and a free 3’ hydroxyl group at the other. The 3’
and 5’ carbons of the sugars are used in a describing the direction a polynucleotide strands runs in a molecule.
WATSON AND CRICK MODEL OF DNA
A model is a visual image of an object or idea which simplifies the object or idea. James Watson and Francis Crick
assembled the molecular model and published their two pahe article on their molecular model of DNA in the journal
“Nature” in Aprail 1953, Few milestones in the history of biology have had a broad an impact as their double helix. Watson
and Crick were awarded Nobel prize in 1962 for their model of DNA.
The important points of this ladder model of DNA are:
1. There are two polynucleotide strands running in apposite directions and winding out each other in a form of double
helix.
2. The double helix looks like a ladder.
3. The sugar and phosphare part of the nucleotide makes the upright part of the ladder.
4. The nitrogen bases of the nucleotide make up the rungs of the ladder.
5. A double ring base purine must always be paired with a single ringed base pyrimidines on the opposite strand
Individual structures of the bases from the pairing more specifically. Each base has chemical side groups that can
best form hydrogen bonds with one appropriate partner.
6. the matching of the basis are specific. Adenine makes the pair with thymine and Gaunine with Cytosine.
7. The base pairs are held together by the hydrogen bond. There are three hydrogen bonds between Guanine and
Cytosine and two hydrogen bonds between Adenine and Thymine.
8. The ratio of A:T and C:G is equal but the ratio of AT:CG is different.
9. The helix is 2nm in diameter and makes a full spiral turn every 3.4 nm i.e. after every ten base pairs. The distance
between tow base pair is 0.34nm.
10. There are two grooves a major grove and a minor grove.
11. The helix rotates about its axis 3.60O longitudinally with each base pair or rug, the spiral winds therough a full
circle i.e. 360O in the length of helix occupied by ten base pairs.
12. There is no restriction on the sequence of nucleotides along the length of a DNA strand. The sequence can vary in
countless ways. The sequence is specific for different species, organisms and even individuals.
DNA Replication
Ability to be replicated is one of the key requirement of a genetic material. Watson and Crick model of DNA
immediately suggested that DNA is replicated by means of complementary base pairing. During replication each old DNA
strand of the parent molecule serves as template for a new strand in a daughter molecule.
DNA replication is termed semi conservative replication because one of the old strands is conserved or is present, in
each daughter molecule Replication requires the following steps.
(CHROMOSOMES AND DNA)
4
Unwinding
The old strands that make up the parent DNA molecule are unwound and “unzipped” i.e. the weak hydrogen bonds
between the paired bases are broken. There is a special enzyme called helicase that unwinds the molecule.
Complementary Base Pairing
New complementary nucleotides always present in the nucleus, are positioned by the process of complementary
base pairing.
Joining: The complementary nucleotides join to form new strands. Each daughter DNA molecule contains an old stand and
new strand.
Complementary base pairing and joining are carried out by an enzyme complex called DNA polymerase.
Meselson and Stahl’s DNA Replication Experiment
Semiconservative replication was experimentally confirmed by Matthew Meselson and Franklin Stahl of Califrnia
Institute of Technology in 1958. Centriguges spin tubes and in that way separate particles from the suspending fluid.
Meselson and Stahl knew that it would be possible to centrifuge DNA molecules in a suspending fluid that would separate
them on the basis of their different densities. A DNA molecule in which both strands contained heavy nitrogen ( 15N), with an
atomic weight 15) is most dense. A DNA molecule in which both strands contained light nitrogen ( 14N, with an atomic
weight of 14) is least dense. A hybrid DNA molecule in which one strand is heavy and one is light has an intermediate
density. Meselson and Stahl first greq bacteria in medium containing 15N so that only heavy DNA molecules were present in
the cells. Then they switched the bacteria to a, medium containing 14N. After one division, only hybrid DNA molecules were
in the cells. After two divisions, half of the DNA molecules were light and half were hybrid. These were exactly the results
to be expected if DNA replication is semicnsevative.
Process of Replication of DNA
Opposite orientation of DNA strand: DNA’s sugar phosphate backbone run in opposite directions. Each strand has a 3’
(“Three prime”) end and a 5’ end. The primed number refers to the carbon atoms of the nucleotide sugar. At one end of each
DNA strand, the sugar’s 3’ carbon atom is attached to an ---OH group, at the other end, the sugar’s 5’ carbon has phosphate
group.
Replication is in 5’ – 3’ direction: The opposite orientation of the strand is important in DNA replication. The enzymes that
link DNA nucleotides replication. The enzymes that link DNA nucleotides to a growing daughter strand, called DNA
polymerase add nucleotide only to the 3’ end of the strand, never to the 5’ end. Thus, a daughter DNA strand can only grow
in the 5’ – 3’ direction.
Replication fork: At the replication fork, DNA is unwound and unzipped and replication occurs.
Leading strand:At the replication fork, only one of the new (daughter) strands runs in the 5’ – 3’ direction. The template for
this strand, of course, runs in the 3’ – 5’ direction. The new strand running in the 5’ – 3’ direction can be synthesized
continuously and is called the leading strand. Thus, the replication is a continuous process for the leading strand.
Lagging strand: But what about the situation when the 5’ – 3’ parental strand is serving as the template? Synthesis of the
new strand must be in the 5’-3’ direction, and therefore synthesis has to begin at the fork. i.e. out2ward from the forking
point. Although synthesis is in the 5’-3’ direction, this new daughter strand in the end runs from 5’-3’ opposite to its
template.
The new strand is synthesized in short segments as the fork opens up. The short segments are called Okazaki
fragments, name after Japanese scientist who discovered them. These fragments are 100 to 200 nucleotides long in
eukaryotes and 1000 to 2000 nucleotide in prokaryotes. Thus, the more time than continuous replication, therefore the new
strand in this case is called lagging strand.
RNA primar: DNA polymerase can join a nucleotide to the free 3’ end of another nucleotide. DNA polymerase cannot strat
the synthesis of a new short amount of RNA called RNA primer (through the action of an enzyme known as primase) that is
complementary to the DNA strand being replicated. Now DNA polymerase can add DNA nucleotides in the 5’ – 3’
direction. Later while prof-reding, DNA polymerase then removes the RNA plymer and replaces it with complementary
DNA nucleotides.
DNA ligase: Another enzyme called DNA ligase, joins the 3’ end of each fragment to the 5’ end of another. When the DNA
is further unwound, new RNA primers are constructed, and DNA polymerase then jumps ahead 1000----2000 nucleotides,
toward the replication fork to begin making another Okazaki fragment.
DNA Replicates in Prokaryotes
Bacteria have a single circular loop of DNA, that must be replicated before cells divides. In some circular DNA
molecules, replication moves around the DNA molecule in one direction only. There are three DNA polymerases namely I,
II and III in bacteria. DNA polymerase I is a relatively small enzyme that plays a supporting role in DNA replication. The
true E. coli replicating enzyme polymerase III is a diamer and catalyzes replication of one DNA strand. Polymerase II
progressisvely threads the DNA through the enzyme complex, moving at a rapid rate, some 1000 nucleotide/ second. One of
the features of the DNA polymerase II is that it can add nucleotide only to a chain of nucleotide that is already paired with
the parent strands. In others, replication starts at the origin byt moves in opposite directions. The process always occurs in
the 5’ to 3’ direction.
DNA Repicates in Eukaryotes
DNA replication begins at numerous origins of replication along the length of the chromosomes, an the so-called
replication bubbles spread bidirectionally. Thousands of bubbles can be present at once. Eventually all the bubbles merge,
yielding two complete daughter DNA.
Errors in DNA replication
A mismatched nucleotide slips through this selection process, only once per 1000,000 base pairs at most. This
mismatched nucleotide cuase a pause in replication, during time, it is excised from the daughter strands and replaced with
correct nucleotide. After this so called proof reading has occurred, the error rate is only one mistake per billion base pairs.
The errors hat slip through the nucleotide selection and proof reading may cause a gene mutation to occur.
What is a gene?
(CHROMOSOMES AND DNA)
5
A unit of inheritance: Medel proposed in 1866 that the characteristics of organisms were determined by hereditary units
which he called elements. These were later termed genes and show to be located on chromosomes, which transmitted them
form generation to generation. Thus a gene may be defined as unit of biological inheritance. This is perfectly acceptable
definition but it does not tell us anything about the physical nature of the gene. The possible ways of overcoming this
objection are considered below.
A unit of recombination: From his studies of chromosomes mapping in Drosophila Morgan postulated that “a gene” was
the shortest segment of chromosme which could be separated from adjacent segments by crossing-over. This definition
regarding the gene as a specific region of the chromosome determine a distinct characteristic in the organism.
A unit of function: A gene can be defined as a place of DNA which codes for protein or more precisely a gene is the DNA
code for a polupeptide, since some proteins are made up of more than one polypeptide chain and therefore coded for more
than one gene.
A unit of gene nutatuon: The change of base during replication causes change in the gene as a result there is change in the
proteins structure and function known as gene mutation.
Genes and Enzymes
Many types of genes function by providing coded information that directs the synthesis of enzymes. Enzymes are
catalysts involved in virtually all biochemical reactions in living cells. If any one of the needed enzyme is not mad, the
particular reaction will not take place, and the cell is unable to function properly.
The discovery of the relationship that exists between genes and enzymes has an interesting history. This
relationship was first suggested by the English physician Archiblad Garrod who studied human metabokic disease. Garrod
and willam Bateson reported in 1902 that ertain findings on the disease alkaptonuria. The individuals with alkaptonuria
produce urine that turns black whicn exposed to air. Garrod observed that the patients excreted homogentisic acid in their
urine. Normal, healthy persons can metabolize homogentisic acid. Garrod hypothesized that people with alkaptonuria lack an
enzyme necessary to catalyze the conversion of homogentisic acid to another compound. Because the disease is inherited in
a simple Mendelian fasion and appeared to be controlled by a single recessive gene. Garrod proposed that there is a direct
relationship between a gene and enzyme.
Experiment of Beadle and Tatum on Neurospora
Neurospora crassa is the red bread mold. It is a haploid organism. It has a single gene for each trait. So there would
be no masking of mutant gene by an allelic partnr. It can reproduce asexually by spores. In sexual reproduction the zygote is
diploid. It is formed in large sac called ascus. The zygote divides into four spores by meiosis. Each spore divides into two by
mitosis. As the spores are formed in the ascus so the spores are called ascospores.
Minimal medium: the medium that contains only a few nutrients i.e. sugar, nitrogen compound, some minerals, salts and
vitamin biotin, on which Neurospora grow is called minimum medium. The ed bread mold can synthesize all the required
amino acids and enzymes form the minimal medium. The neurospora that can grow in the minimal medium is called wild
type.
Mutant: When there is irradiation with X-rays on the mold, mutation occurs. The molds are called nutant molds. The
mutants do not grow on the minimal medium.
Biochemical medium: The medium that contains the amino acids or vitamins for the growth of the mutant mold is called
biochemical medium or complete medium.
Experiments
In 1951 G.W. Boadle and E.Z. Tatum conducted a series of experiments on Neurospora crassa. They used X rays to
induce mutatuin in the mold. Fig. 20.21 illustrated the experiment. In this experiment spores were first irradiated to induce
mutation and were placed on complete medium and allowed to grow. Once the colonies were established, individual spores
were taken and tested to see whether they would grow on minimal medium which lacks the amino acids and vitamins that
the fungus normally manufactures. Any strains that will not grow on minimal medium but will grow on complete medium,
contains one or more mutations in the genes that are necessary to produce one of the substances in the complete but not in
the minimal medium. To find out any particular mutation a single spore was selected and was placed in the compete
medium. The colonies were extablished. A spore was transfeered on to minimal medium. Then the different test tube of
minimal medium was supplemented with one particular substance. The spore grew on minimal medium to which only
arginine was substance. The spore grew on minimal medium to which arginine was added. It was called arg (arginine)
mutant. When the chromosomal position of each mutan arg was located, they were found cluster in three areas.
For each enzyme in the arginine biosynthetic pathway, Beadle and Tatum were able to isolate a nutant strain with a
defective form of that enzyme and the mutation always proved to be located at one of a few specific chromosomal sites, a
different site for each enzyme. The gene produce their effects by changing the structures of enzymes i.e. each nutatuin alters
a single gene that controls one step in the synthesis of a particular kind of mutation. The geneticists called this relationship
the one gene one enzyme hypothesis.
This series of compounds-orinithine, citrulline and arginine, whose story had been worked out in Neurospora also
occurs in amny other species including man.
One Gene One Polypeptide hypothesis
The one gene one enzyme mutation causes a change in the structure of a protein. Proteins are polymers of amino
acids, some of which carry a charge. Linus Pauling and Harvey ltano decided to see if there was a charge difference between
normal haemoglobin (HbA) and sickle cell hemoglobin (Hb S). To determine this, they subjected haemoglobin collected from
sickle cell trait individuals and sickle-cell disease individuals to electrophoiresis (fig no. 23.9) a procedure that separates
molecules according to their size and charge, whether (+) or (-). Here is what they found. As you can see in the fig. 22.23,
there is a difference in migration rate towards the positive pole between haemoglobin and sicikle cell hemoglobin. Further,
haemoglobin from those with sickle-cell trait separates into two distinct bands, one corresponding to that of HbA
haemoglobin and other corresponding to that HbS haemoglobin. Pauling demonstrated that a mutation leads to a change in
the structure of protein.
(CHROMOSOMES AND DNA)
6
Fredirick Sange (1953) described for the first time that insulin consists of definable sequence of amino acids.
Enzymes and other proteins consist of chain of amono acids arranged in certain definite order. In 1956 Veron Ingram was
able to determine the structural difference between HbA contains negatively charged glutamate at 6th postion, in sickle cell
haemoglobin the glutamate is replaced by nonpolat valine. Haemoglobin contains two types of polypeptide chains, designed
α and β. Only the beta chain is affected in persons with sickle-cell trait and sickle cell disease, therefore, there must be a
gene for each type of chain. A refinement of he one gene one enzyme hypothesis was needed, and it was replaced by the one
gene one polypeptide hypothesis, i.e. a gene can change the structure and function of polypeptide chain.
FLOW OF GENETIC INFORMATION IN CELLS
From DNA to Proteins
DNA is a linear polymer of nucleotides and polypeptides are linear polymers of amino acids. This colinearity
suggests that the nucleotide suqence of DNA somehow determines the order of amino acids in proteins. A DNA molecule,
however, cannot directly control the sequence of amino acids because DNA is found in the nucleus of eukaryotes (or the
nucleiod in prokaryotes), vand plrotein synthesis olccurs in the cytoplasm. The molecules that act as a bridge between, is
RNA found in both the nucleus and the cytoplas,.
RNA
RNA is a polymer of nucleotiede. It consists of sugar ribose and the bases Adenine (A), Cytosine (C), Guanine (G),
Uracil (U). RNA is a single stranded and does not form a double helix in the same manner as DNA. There are three major
classes of RNA, each with special functions in protein synthesis:
Messenger RNA (mRNA): It takes a message from DNA in the nucleus to th ribosomes in the cytoplasm.
Ribosomal RNA (rRNA): Along with proteins, it makes up the ribosomes, where proteins are synthesized.
Transfer RNA (tRNA):It transfers amino acids to the ribosomes.
The process by which an RNA is made according to the base sequence of a portion of DNA is called transcription.
The process by which an mRNA transcri-ption direct the sequence of amino acids in a polypeptide is called translation.
Transcription:
The molecules mRNA, rRNA and tRNA is transcribed from DNA template. In prokaryotes there is only one type of
RNA polymerase for the synthesis of RNA. In eukaryotes there are three types of RNA polymerase for the synthesis of three
types of RNA i.e. polymerase I (rRNA), polymerase II (mRNA), polymerase III (tRNA).
mRNA is formed: Transcription begins when RNA polymerase II attaches to a region of DNA called promoter. A promoter
defines the start of a gene, the direction of transcription, and the strand to be copied. Once RNA polymerase is attached the
DNA helix opens. One of the two strand of DNA which is to be transcribed is called template or the antisense strand. The
opposite strand is called coding strand or the sense strand.
In prokaryotes within promoter there are two binding sites TTGACA also called 35 sequence and TATAAT
sequence also called 10 sequence which have affinity for RNA polymerase. One of the subunits of RNA transcription
process. Once the transcription has started the sigma factor is released. RNA polymerase proceeds to read DNA bnase
sequence as a template for assembly of a complementary base in RNA molecue. The RNA polymerase synthesize from 5’ to
3’ direction. RNA polymerase add one nucleotide at a time to form the RNA molecule. When it meets a terminal signal, a
specific sequence in DNA that signals the termination transcription. The simplest termination or stop singnal is a series of
GC base pairs followed by a series of AT base pairs. The RNA formed in this region forms a GC hair-pin followed by four
U ribonucleotides. The hairpin causes RNA polymerase to stop synthesis. In bacteria the newly synthesized mRNA is
directly released into the cytoplasm.
mRNA is processed: The newly formed mRNA, called primary mRNA transcript is eukaryotic nucleus. (1) the ends of he
mRNA molecule are altered: a cap is put onto the 5’ end a “poly-A tail” is put onto the 3’ end. The cap is a midified guanine
(G) nucleotide that helps to tell a ribosome where attach when translation begins. The “poly-A tail consists of a chain of 150
to 200 adenine (A) nucleotide. The caps and tails save the mRNA from variety of nuceases and phosphateses, so that the
molecule reain stable during long journey to ribosomes. (2) Portions of the primary mRNA transcript are removed. The
portions that remain are called exons because they will be expressed. In between the exons are intervening segments called
introns that are removed during an mRNA processing event called mRNA splicing. The mRNA that has now been processed
and leaves the nucleus is called mature mRNA. Once the portionof the DNA molecule is transcribed the double helix
reforms.
rRNA is formed: It is transcribed bt the genes present.on the DNA of the several chromosomes found within the region of
the nucleolus known as nuclear organizer. The enzyme involved is polymerase I. It is called rRNA because it eventually
becomes part of ribosomes. The rRNA is packaged with a variety of proteins into ribosomal subunits, one of which is large
than the other. The base sequence of rRNA is similar form bacteria to higher animals and plants.
tRNA is formed: It is also transcribed as per DNA template RNA polymerase III synthesizes tRNA. It is the smallest of the
RNA molecule. A tRNA is a single-stranded nucleic and folds double on itself to create regions where complementary bases
are bounded to one another. The structure of a tRNA molecule is generally known as a flat colover leaf.
The whole molecule consists of 80 nucleotides byut only 20 shows the complementary base pairing. There are at
least one tRNA molecule for each of the 20 amino acids found in proteins. Sixty tRNA have been identified.
Human cells contain about 45 different kinds of tRNA molecules. The 5’ end, ends in Guanine base while the 3’
end always is the base sequenced of ACC. The nucleotide sequence of the rest of the molecule is variable. tRNA middle
three of which form the anticodon, it is complementary to specific condon of mRNA. For example, a tRNA that has
anticodon GAA binds to the condon CUU and carries amino acid leucin. The D loop recognizes the activation enzyme.
Theat (θ) loop recognizes the specific place on the ribosome for binding during protein synthesis.
Genetic code
The sequence of nucleotide in DNA specifies (and a copy of this sequence in mRNA directs) the order of amono
acids in a polypeptide. This relation between the amino acid and the bases is called genetic code. It can four nucleotide
provide enough combinations to code for 20 amino acids? If each code word called a codon, where made of one base, the
(CHROMOSOMES AND DNA)
7
protein could containonly 4 amino acids. If the combination of two base codes for be specified into protein molecules. But of
each condon were made up of three bases there would be 64 condons more than enough to code for 20 amino acids.
Finding the genetic Code
In 1961, Marchall Nirenbery and J. Heinrich Matthei constructed synthetic RNA which composed only of Uracil,
and the protein that resulted was compsed of only amino acid phenyulalanine. Therefore a condon for phenylalanine was
known to be UUU. Later a cell free system was developed by Nirenberg and Philip Leder in which three nucleotides at a
time were translated; in that way it was possible to assign an amino acids to each of the RNA codons.
Important Properties of Condon
Degenerate: The genetic code is “degenerate”. This means that most amino acids have more then one codon.
Unambiguous: The genetic code is unambiguous.. Each triplet codon has only one meaning.
Initiation codon: There is only one initiation codon which is AUG.
Punctuation codon: There are three srop signal. These are UAA, UAG, UGA.
Universal triplet of bases: The tripet code for an amino acids is universal in all organisms.
Non overlapping: mRNA sequence beginning AUGAGCGCA is not read as AUG/ UGA/GAG. It will be read as
AUG / AGC / GCA.
Protein synthesis
The two main steps of protein synthesis is transcription and translation.
Transcription
The genetic message is in DNA. From DNA template a single strand of mRNA is produced. This process is called
transcription (which we have already discussed, see fig. 20.26). It is called mRNA because it carries the genetic message
form DNA. The triplet of DNA is called code and the tripet of mRNA is called codon.
Translation:
It is the mechanism by which the triplet base of mRNA forms specific sequence of amino acids in a polypeptide
chain. The main steps involved in the translation are:
(1) Binding mRNA with the ribosome
(2) Amino acid activation
(3) Attachment of amino acid to tRNA
(4) Polypeptide chain initiation
(5) Chain elongation
(6) Chain termination
Binding of mRNA to Ribosome
The mRNA attaches itself to a 30S ribosome. The 50S subunit joins then 30S to form 80S ribosome. On a fully
assembled ribosome there are two sites.
2. Amino Acid Activation
For each amino acid there is an enzyme. With the presence of this enzyme the amino acid becomes attached with
AMP and then amino acid is activated. The enzyme is called activation enzyme.
ATP + Amino Acid + Enzyme = Enzyme – Amino Acid – AMP +PP
3. Attachment of Amino Acid to tRNA
The D loop of the tRNA recognizes the activation enzymes. An amino acid becomes attached to a particular tRNA
at the 3’ end i.e. A-C-C- end, by its carboxyl group.
4. Chain Initiation
As we have already seen that a ribosome has two binding sites for tRNAs. One of these is called P (for peptide) site
and the other is called the A (for amino acid) site. The anticodon UAC of tRNA matches with the codon AUG. The ribosome
exposes the condon on the mRNA immediately adjacent to the initiating AUG condon. The tRNA where anticondon will
match with this condon will become attached with the ribosome at A site. The two amino acids will react and form the
peptide bond. The methionine tRNA will be released.
5. Chain Elogation
In the process of elongation ribosome will move along the mRNA molecule a distance corresponding to the three
nucleotide. Another tRNA carrying a specific amino acid links up with condon. The amino acid becomes linked to the
previous amino acid and tRNA at site P is released. The process of ribosomal movement continues along the mRNA and a
polypeptide is fomed.
6. Chain Terminiation
When the ribosome comes to a condon signaling “stop”, the joining of condon. The teminating codon are UAA,
UAG and UGA. At this stage polypeptide chain leaves, the ribosme, and the translation is completed.
Muatations
Defination
Mutations (L. Mutatus, change) are permanent changes in genes or chromosomes that can be passed to offspring if
they occur in cells that become gametes. There are two main types of mutation:
(1) Chrmosomal Mutations (2) Gene mutations
Chrmosomal Mutations
The chromosomal mutation include:
(a) Changes in chromosome number
(b) Changes in chrmosme structure
Changes in Chromosome Number:
Changes in chromosome number include monosomy, trisomy, and poluyploidy.
Monosomy and Trisomy
Monosomy occurs when an individual has only one of a particular type of chrmosme (2n-1) and trisomy occurs
when an individual has three of a partifular type of chromosme (2n+1). The usual cause of monosomy and trisomy is
(CHROMOSOMES AND DNA)
8
nondisjuction during meiosis. Monosomy and trisomy in both plants and animals. In human Turner syndrome is a
monosomy of the sex chromosomes, the individual inherit a single X chromosome. The most common trisony among human
is Down syndrome, which involves chromosme 21. You will study about these in the next chapters.
Polyploidy
Some mutansts have more than two sets of chromosomes. They are called polyploids. Polyploid organism may be
triploid (3n), tetraploid (4n) pentaploids (5n) and so on. Polyploidy is not seen in animals. It is estimated that 47% of all
flowering plants are polyploids. Crops such as wheat, corn, contton, sugar cane and fruits such as watermelons, bananas and
apples are polyplodis. Polyploids generally arises by following hybridization.
Changes in Chromosme Structure
There are various agents in the environment, such as radiation, certain organic chemicals or even viruses that can
cause chromosme to break. When the broken ends of chromosomes do not rehoin in the same pattern as before this results in
a change in chromosomal structure.
Inversion: It occurs when a segment of a chromosome is turned around 180 O.
Translocation: It is the movement of a chromosome segment from one chromosme to another non homologous chromosme.
Deletion: It occurs when an end of a chromosme breaks off or when two simultaneous breaks lead to the loss of an internal
segment. An example is cri du chat (cat;s cry) syndrome.
Duplication: It is the doubling of chromosme segment.
Gene Mutations
Gene mutation is a change in the nucleotide sequence of DNA. If this sequence changes, then the condons change
and sequence of amino acids in a polypeptide changes.
Frame shift mutations:
The mutations occur most often because one or more nucleotides are either inserted or deleted from DNA. This results in a
completely new sequence of condons and a nonfunctional protein.
Point nutations: These involve a change in a single nucleotide and therefore a change in a specific condon. The occurrence
of valine instead of glutamate in the β (beta) cahin of haemoglobin (hemoglobin) results in sickle cell disease. Due to point
mutation there is exzyme deficiency and homogentisic acid is nit further changed into 4 maleyacetoacetic acid as this leads
to alkaptonuria.
EXERCISE – SECTION – I – OBJECTIVE QUESTIONS
1. Fill in the blanks.
(i). The whole characteristics of chromosomes can be studied by its ___________.
(ii). The nucleoprotein bead is a ___________.
(iii). The helical DNA is about ______ in diameter.
(iv). The replication of DNA is _________ in nature
(v). The lagging strand of DNA is synthesized by short fragments called ________
(vi). When glutamic acid replaced by valine the result is ____
(vii). During protein synthesis _____ provides the site for the attachment of coming amino acids.
(viii). The password for specific amino acid is in the form of _______.
(ix). Down syndrome is a genetic disease that is due to ________.
(x). The initiation codon is ____________.
2. Mark the statements as “True” or “False”.
(i). The sister chromatids are attached at a point called centromere.
(ii). DNA along with its protein make a complex called nuycleotide.
(iii). The DNA replication starts only one site.
(iv). The enzyme that catalyze DNA replication is a polymer.
(v). The changes in DNA that results in abnormalities are called Mutation.
(vi). Highly condensed portion of chromotin is called heterochromatin.
(vii). In alkaptonuria the urine of patients contain glucose in excess.
(viii). Promotors are used to stop the process of transcription.
(ix). In Prokaryotes three types of RNA synthesized are the mRNA, rRNA, tRNA.
(x). Insertion is an abnormality due to point mutation.
3. Select the correct answer and encircle it.
(i). Which structure becomes visible when cell starts dividing (a) nucleus (b) cell membrane (c) chromosomes (d)
nuclear membrane
(ii). The bond exists between N2 bases of DNA is: (a) hydrogen (b) covalent (c) ionic (d) phosphodiester
(iii). Attachment of Okazaki fragments to DNA’s lagging strand is facilitated by: (a) ligase (b) polymerase II (c)
polymerase III (d) DNA ase
(iv). Formation of RNA from DNA is process called (a) transcription (b) translation (c) replication (d) none.
(v). UAA represents: (a) stop condon (b) nonsence condon (c) promoters (d) both a and b
(vi). The caps and tails attached to mRNA protect it from: (a) polymerases (b) none (c) ligases (d) nucleases and
phosphateses
(vii). Vernon Ingram discovered molecular basis of: (a) sickle cell anemia (b) cancer (c) phenylketonuria (d) all
(viii). The strand of DNA being transcribed is called template or: (a) gene mutation (b) stop mutation (c) point mutation
(d) none
(ix). The replication of DNA is always (a) 5-3 (b)both (c)3-5 (d) none
4. Match the following columns.
(i) Column ‘A’ Column ‘B’
a. Genes i. DNA model
(CHROMOSOMES AND DNA)
9
b. Wirulent S. pneumoniae ii. DNA polymerases
c. Watson and Crick iii. Meselson and Stahl
d. DNA replication iv Locus
v. S form
(ii) Column ‘A’ Column ‘B’
a. Initiation of transcription i. GC+AT base point
b. Stop signal ii. DNA replication
c. Poly A-tail iii. U ribonucleotides
d. Transcription bubble iv Sigma factor
v. 3’ end
(iii) Column ‘A’ Column ‘B’
a. Alkaptonuria i. Chromosmal aberration
b. Inversion ii. 1000 nucleotides/sec
c. P-site iii. Homogentisic acid
d. Polymerase III iv Peptidyl site
v. ACC
5. Draw and label a tRNA
SECTION – II – SHORT QUESTIONS
1. Define the following:
karyotype nucleosome euchromatin
heterochromation purine pyrimidines
nucleoside nucleotide phosphodiester dond
base okazake fragment gene nutation
template code codon
anticodon transcription translation
molecular biology
2. Differentiate between:
Karal correns
Walter Sutton
Frederick Griffith
Alfred Hershy
Martha Chase
Waston
Crick \
Rosalind Franklin
Mauric wikins
Erwin Chargaff
Archibald Garrod
Beadle and Tatum
Frderick Sanger
Vernon lngram
Hofmeister.
Walther Flemming
Waldyer
Oswald Avery
3. Differentiate between:
DNA and RNA
Code, codon and anticodon.
Monomer and polymer.
Purine and pyrimidine.
Nucleosome and nucleotide
Transcription and translation
Euchromatin and heterochromatin
4. Name the types of chromosme on the basis of location of centromere.
5. The base sequence of one strand of a DNA molecule is: G-A-C-G-T-A-C. What is the sequence of the bases of the
other strand?
6. Writre the base sequence of mRNA from a DNA strand with the following sequence: A-T-G-T-T-C-G-A-G-T-A-C-
C-A-T-G
7. How an amino acid is activated during protein synthesis?
8. Name the main steps of protein synthesis.
SECTION – III – EXTENSIVE QUESTIONS
1. Describe chemical composition and coiling of DNA
2. Describe the structure and replication of DNA.
3. What are chromosomes? Describe the types of chromosomes and chromosomal mutation.
4. Discuss chromosme theory of inheritance.
(CHROMOSOMES AND DNA)
10
5. How it was confirmed that DNA is the hereditary material?
6. Give an account of Genetic Code.
7. What is a gene? Discuss one gene one enzyme hypothesis and one gene one polypeptide hypothesis and gene mutation.
8. Describe the structure of three types of RNA
9. Describe the process of translation.
10. Describe protein synthesis.
ANSWERS
1.
(i) Karyotype
(ii) nucleosome
(iii) 2nm
(iv) semi conservatiove
(v) Okazaki fragments
(vi) Sickle cell anemia
(vii) rRNA
(viii) Genetic code
(ix) Chromosomal
(x) Nothing
2.
(i) T
(ii) F
(iii) F
(iv) T
(v) F
3.
(i) c
(ii) a
(iii) a
(iv) a
(v) d
(vi) d
(vii) a
(viii) c
(ix) c
(x) a
4.
(i) (a) iiv (b) v (c) i (d) ii
(ii) (a) iv (b) i (c) v (d) ii
(iii) (a) iii (b) i (c) iv (d) ii