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Transcript
DNA AS GENETIC MATERIAL,
CHEMISTRY OF NUCLEIC
ACIDS
AND GENETIC CODE
DNA
• The structure of DNA encodes all the information
every cell needs to function and thrive.
• DNA carries hereditary information in a form that
can be copied and passed intact from generation
to generation.
GENE
• A gene is a segment of DNA.
• The biochemical instructions found within most
genes, known as the genetic code, specify the
chemical structure of a particular protein.
• Proteins are composed of long chains of amino
acids, and the specific sequence of these amino
acids dictates the function of each protein.
Evidence that genes are made of
DNA (or sometimes RNA)
1. Transformation in Bacteria
• Frederick Griffith laid the foundation for the identification
of DNA as the genetic material in 1928 with his
experiments on transformation in the bacterium
Pnuemococcus, now known as Streptococcus pneumoniae.
2. DNA: The transforming material.
• Oswald Avery, Colin Mac Leod, and Maclyn Mc Carty
showed the transforming substance to be DNA in 1944 in
virulent cells of Streptococcus pneumoniae.
• In 1952, A.D. Hershey and Martha Chase performed
experiment in T2 bacteriophage. The phage is composed of
protein and DNA only. The experiment showed that the
genes of phage are made of DNA.
The chemical nature of Polynucleotides
• By the mid 1940s, biochemists know the fundamental
chemical structures of DNA and RNA.
• When they broke DNA into its component parts, they
found these constituents to be nitrogenous bases,
phosphoric acid, and the sugar deoxyribose.
• RNA yielded bases and phosphoric acid, plus a
different sugar ribose.
• The four bases found in DNA are adenine (A),
cytosine (C), guanine (G) and thymine (T).
• RNA contains the same bases, except that Uracil (U)
replaces thymine.
• Adenine and Guanine are purines and are two ringed
structures.
• Thymine,Cytosine and Uracil are single ringed and
are called pyrimidines.
• These structures constitute the alphabet of genetics.
Ribose contains a hydroxyl (OH) group in the 2 –
position.
• The subunits of DNA and RNA are nucleotides,
which are nucleosides with a phosphate group
attached through a phosphodiester bond.
DNA Structure
• Linus Pauling elucidated that the  - helix, an important
feature of protein structure.
• Maurice Wilkins and Rosalind Franklin, another group
tried to find out the structure of DNA at Kings College in
London. They used X-ray diffraction to analyse the threedimensional structure of DNA.
• Erwin Chargaff was another very important contributor.
Chargaff studies (1950) of the base composition of DNAs
from various sources revealed that the content of purines
always equaled the content of pyrimidines.
• the amounts of adenine and thymine were always equal, as
were the amounts of guanine and cytosine. These findings,
known as Chargaff's rules, provided a valuable
confirmation of Watson and Crick's model.
• Franklin's X-ray work strongly suggested that DNA was a
helix.
The Double helix
• DNA molecules form chains of building blocks called
nucleotides.
• Each nucleotide consists of a sugar molecule called
deoxyribose that bonds to a phosphate molecule and to a
nitrogen-containing compound, known as a base.
• DNA uses four bases in its structure: adenine (A), cytosine
(C), guanine (G), and thymine (T).
• The order of the bases in a DNA molecule—the genetic
code—determines the amino acid sequence of a protein.
• In the cells of most organisms, two long strands of DNA join
in a single molecule that resembles a spiraling ladder,
commonly called a double helix.
• Alternating phosphate and sugar molecules form each side of
this ladder.
COMPLEMENTARY BASE PAIRING
• Adenine always joins with Thymine
Guanine always links to Cytosine.
• Scientists use complementary base pairing to help identify the
genes on a particular chromosome and to develop methods used
in genetic engineering.
• Watson and Crick found that the best model that satisfied all the
X-ray data was a double helix
• The two chains run in an anti parallel fashion with one chain
having a 51  31 orientation and the other having a 31  51
orientation.
• The width of the helix was found to be 2 nm.
• The purine and pyrimidine bases were stacked 0.34 nm apart in
a ladder.
• The helix made one full turn every 3.4 nm and, therefore, there
should be 10 layers of bases stacked in one turn..
• In a given DNA, adenine is equal to thymine and guanine to
cytosine.
• There are two hydrogen bonds for A = T pairing and three
bonds for C  G pairing.
• C  G pairing is more stronger than A = T pairing.
• Helical structure is right handed.
• The fifth (5- prime, of 5') carbon of the pentose ring is
connected to the third (3 - prime, of 3 ') carbon of the next
pentose ring via a phosphate group, and the nitrogenous bases
stick out from this sugar-phosphate back bone.
• By convention, DNA sequences are read from 5'→3' with
respect to the polarity of the strand.
Genes made of RNA
• A group of viruses, referred to as retroviruses, has
RNA as the genetic material.
• These tumour viruses can integrate with the host
genome DNA, only after the RNA makes a DNA
copy.
• The central dogma says that the flow of information is
unidirectional .i.e. ,DNA → RNA → Protein.
• With the discovery of the enzyme reverse
transcriptase, RNA can also go back to DNA and the
central dogma is now represented as:
RNA → DNA → Protein.
A variety of DNA structures
• B form is present in most DNA in the cell. The plane
of a base pair is no longer perpendicular to the
helical axis, but tilts 20 degrees away from
horizontal.
• The A helix packs in 11 base pairs per helical turn
instead of 10 found in the B form, and turn occurs in
31 angstroms instead of 34.
• The distance between base pairs, is only 2.8 nm
instead of 3.4 nm, as in B-DNA.
• Both the A and B form DNA structures are right
handed; the helix turns clockwise.
• Alexander Rich and his colleagues discovered in
1979, DNA can exist in an extended left-handed
helical form
• The zigzag look of this DNA's backbone when
viewed from the side, it is often called Z DNA.
• The distance between base pair is 4.5 nm and
number of bases per turn is 12.
• RNA-DNA hybrid strand assumes the A form.
• Normal DNA has 2 groove (major and minor).
• Z- DNA has single groove.
Separating the two strands of a DNA double helix
• While the ratios of G to C and A to T in an organisms
DNA are fixed,
• the GC content (percentage of G + C) can vary
considerably from one DNA to another.
• The values of GC content range from 22% to 73%
and these differences are reflected in differences in
the properties of DNA.
PROPERTIES OF DNA
1. DNA denaturation, or DNA melting
The temperature at which the DNA strands are half denatured is
called the melting temperature, or Tm. This is known as DNA
denaturation.
• The amount of strand separation or melting is measured by the
absorbance of the DNA solution at 260 nm.
• The higher a DNA's GC content, the higher its Tm. C  G
pairing form 3 hydrogen bonds, whereas A = T pairs have
only 2.
• In addition to heating, DMSO and formamide also disrupt
the hydrogen bonding between DNA.
2. Annealing or Renaturation
• Once the two strands of DNA separate, they can under the
proper conditions, come back together again. This is called
annealing or renaturation.
• Factors that contribute to renaturation are
• 1. Temperature : Best temperature for renaturation of a DNA
is about 25oC below its Tm
• 2. DNA concentrations: The higher the concentration, the
faster the annealing
• 3. Renaturation time: If longer time allowed for annealing,
the more will occur.
Activities of genes
• A gene is a unit of information which is held as a
code in a discrete segment of DNA.
• This code specifies the amino acid sequence of a
protein
• The coding parts of a gene sequence are exons, and
the non- coding parts are introns.
• Before a gene can be expressed, the DNA that
encodes has to be transcribed into RNA.
A gene participates in 3 major activities
1. A gene can be replicated
• genetic information can be passed from generation to
generation unchanged.
2. The sequences of bases in the RNA depends directly on the
sequences of bases in the gene.
• Most of these RNAs, in turn, serve as templates for making
protein molecules.
• Thus, most genes are essentially blueprints for making
proteins.
• The production of protein from a DNA blueprint is called gene
expression.
3. A gene can accept occasional changes, or mutations.
GENETIC CODE
• Most genes encode proteins and although only a small part of
the total DNA ,the coding regions of genes act as a template
for the protein.
• Proteins are made up of amino acids.
• It is the sequence of amino acids which give the protein its
specific properties.
• DNA template is first transcribed into mRNA.
• The mRNA template is then translated into a chain of amino
acids.
• There are 20 different amino acids which are used to build up
proteins.
• Marshall Nirenberg and his associates in the early
1960sdevised an elegant technique, called the triplet
binding test, and discovered the first word of code
dictionary.
• A system was developed for synthesizing proteins in
vitro.
• The system included a cell extract containing
ribosomes, tRNAs and other cellular components.
• When synthetic mRNAs consisting entirely of a single
type of nucleotide were added, polypeptides composed
of only a single type of amino acid were formed.
• Thus phenylalanine was formed when polyuridylic
acid (poly U) was added.
• Marshall Nirenberg, Severo Ochoa, Hargobind
Khorana, Francis Crick and many others
contributed significantly to decipher the genetic
code.
• They figured out that the order in which amino
acids are arranged in proteins.
• On the basis of a variety of experiments, it was
found out that a particular sequence of 3 bases
(triplet) would code for a particular amino acid
and this triplet is referred to as codon.
• Many amino acids have more than one codon
and codons specifying the same amino acid are
said to be degenerate and differ in only the
third base.
• The complex process by which the information
in RNA is decoded into a polypeptide is an
exciting story and its understanding represents
a great achievement of the twentieth century in
Biology.
Properties of the Genetic code
• The code is highly degenerate, i.e.,most of the amino
acids are coded for more than one amino acids.
• Leucine, serine and arginine have 6 different codons.
• Proline, threonine and alanine, have four.
• Isoleucine has three.
• Methionine and tryptophan have only one codon.
• The code is not overlapping. There is no punctuation
or spacing between different codons.
• The starting signal for protein synthesis is the codon
AUG (for methionine).
•
• The code appears to be highly universal, i.e.,
it is the same for various different kind of
organisms.
• Coding regions can be transferred from one
organism to another and the correct protein
produced.
• However, a few exceptions to this are known.
For example, in yeast mitochondria, UGA
codes for tryptophan instead of stop.
• In Paramecium, UAA and UAG code for
glutamine instead of stop codon
Point Mutation will cause change in the amino acid sequence
Normal gene frame
Delete 1 base (F)
Add 1 base (X)
1
2
3
4
5
BIG FAT CAT ATE RAT
BIG ATC ATA TER ATBIG FAT CXA TAT ERA +
The universality provides strong evidence that life on earth
started only once.
• When the first living forms appeared some 3 billion years
ago, the genetic code was established and it has not
changed since then through out the evolution of living
organisms.
• The selective pressure has been less strict in mitochondrial
DNA. Mitochondria code only for few proteins and have
their own protein synthetic machinery. The overall code
has been maintained
THANK YOU