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Chemical Structure of Deoxyribonucleic
Acid.
Evidences, DNA is genetic material.
Chromosome and Chromatin.
Anatomy of Chromosome
Dr. Kamal Omer Abdalla,
Associate Prof. of Biochemistry and Molecular
Biology,University of Gadarif.
• Although the name nucleic acid suggests
their location in the nuclei of cells, yet
some of them are, however, also
present in the cytoplasm.
• The nucleic acids are the hereditary
determinants of living organisms. They
are the macromolecules present in most
living cells either in the free state or
bound to proteins as nucleoproteins.
• There are two types of nucleic acids,
deoxyribonucleic acid (DNA) and ribonucleic acid
(RNA). Both are present in all plants and
animals. Viruses also contain nucleic acids,
however, unlike a plant or animal has either RNA
or DNA, but not both.
• DNA is found mainly as a component of
chromatin material of the cell nucleus whereas
most of the RNA (90%) is present in the cell
cytoplasm and the remaining (10%) in the
nucleolus. Extra-nuclear DNA also exists, for e.g.,
in mitochondria and chloroplasts.
Composition of nucleic acids
• Nucleic acids are biopolymers of high molecular
weight with mononucleotide as their repeating
units. Each mononucleotide consists of the
following: 1. Nitrogenous bases 2. Phosphoric
acid and 3. Pentose sugars
Nitrogenous bases
• Two types of major nitrogenous bases, which
account for the base composition of DNA or RNA,
are found in all nucleic acids. These are: a) Purine
bases and b) Pyrimidine bases. The purine and
pyrimidine bases found in nucleic acids are shown
in Fig. 1.
Phosphorus
• Phosphorus, present in the backbone of
nucleic acids, is a constituent of
phosphor-diester bond that links the
two sugar moieties. The molecular
formula of phosphoric acid is H3PO4. It
contains three mono-valent hydroxyl
groups and a divalent oxygen atom, all
linked to the pentavalent phosphorus
atom.
Sugar
• Both DNA and RNA contain five-carbon aldose sugar,
i.e. a pentose sugar. The essential difference
between DNA and RNA is the type of sugar they
contain. RNA contains the sugar D-ribose (hence
called ribonucleic acid, RNA) whereas DNA contains
its derivatives 2’-deoxy-D-ribose, where the 2’hydroxyl group of ribose is replaced by hydrogen
(hence called deoxyribonucleic acid, DNA). Sugars
are always in closed ring β-furanose form in nucleic
acids and hence are called furanose sugars because
of their similarity to the heterocyclic compound
furan.
Why is 2'-Deoxyribose the Sugar Moiety in DNA?
• Common perhydroxylated sugars, such as
glucose and ribose, are formed in nature as
products of the reductive condensation of
carbon dioxide we call photosynthesis. The
formation of deoxysugars requires additional
biological reduction steps, so it is reasonable to
speculate why DNA makes use of the less
common 2'-deoxyribose, when ribose itself
serves well for RNA.
• At least two problems associated
with the extra hydroxyl group in
ribose may be noted. First, the
additional bulk and hydrogen
bonding character of the 2'-OH
interfere with a uniform double
helix structure, preventing the
efficient packing of such a molecule
in the chromosome.
• Second, RNA undergoes spontaneous hydrolytic
cleavage about one hundred times faster than
DNA. This is believed due to intramolecular
attack of the 2'-hydroxyl function on the
neighboring phosphate diester, yielding a 2',3'cyclic phosphate. If stability over the lifetime of
an organism is an essential characteristic of a
gene, then nature's selection of 2'-deoxyribose
for DNA makes sense. The following diagram
illustrates the intramolecular cleavage reaction
in a strand of RNA.
• Structural stability is not a serious challenge
for RNA. The transcripted information
carried by mRNA must be secure for only a
few hours, as it is transported to a
ribosome. Once in the ribosome it is
surrounded by structural and enzymatic
segments that immediately incorporate its
codons for protein synthesis. The tRNA
molecules that carry amino acids to the
ribosome are similarly short lived, and are
in fact continuously recycled by the cellular
chemistry.
• The structural difference in the sugars of DNA
and RNA, though minor, confers very
different chemical and physical properties
upon DNA than RNA.
• RNA is much stiffer due to steric hindrance
and more susceptible to hydrolysis in alkaline
conditions, perhaps explaining in part why
DNA has emerged as the primary genetic
material. Sugar, along with phosphate
performs the structural role in nucleic acids.
Nucleotides or Nucleoside 5’-triphosphates
• These are phosphate esters of nucleosides
i.e. nucleosides form nucleotides by joining
with phosphoric acid. Esterification can occur
at any free hydroxyl group, but is most
common at the 5′and3′positionsin sugars.
The phosphate residues are joined to the
sugar ring by a phosphor monoester bond
and several phosphate groups can be joined
in series by phosphor anhydride bonds.
• These occur either in the free form or as
subunits in nucleic acids. The phosphate is
always esterified to the sugar moiety. The
trivial names of purine nucleosides end with
the suffix –sine, and those of pyrimidine
nucleosides end with suffix –dine.
• In addition to their role as structural
components of nucleic acids, nucleotides also
participate in a number of other functions as
described below:
Energy carriers: Nucleotides represent energy
rich compounds that drive metabolic process,
especially biosynthetic, in all cells. Hydrolysis
of nucleoside triphosphate provides the
chemical energy to drive a wide variety of
cellular reactions. ATP is the most widely
used for this purpose. UTP, GTP, CTP are also
used. Nucleoside triphosphate also serves as
the activated precursors of DNA and RNA
synthesis.
• The hydrolysis of ester linkage (between
ribose and α-phosphate) yields about 14 kJ /
mol under standard conditions, whereas
hydrolysis of each anhydride bond (between
α-β and β-γ phosphates) yields about 30 kJ /
mol. ATP hydrolysis often plays an important
thermodynamic role in biosynthesis.
• Enzyme cofactors: Many enzyme cofactors
include adenosine in their structure, e.g.,
NAD, NADP, FAD.
• Chemical messengers: Some nucleotides act
as regulatory molecules and serve as chemical
signals or secondary messengers, key links in
cellular systems that respond to hormones
and other extracellular stimuli and lead to
adaptive changes in cells interior. Two
hydroxyl groups can be esterified by the same
phosphate moiety to generate a cyclic AMP
(cAMP, adenosine 3’-5’ cyclic phosphate) or
cyclic GMP (cGMP, guanosine 3’-5’ cyclic
phosphate).
Oligonucleotides
• Oligonucleotides are polymers containing <
100nucleotides. These nucleotides are linked
by phosphodiester bond as shown in Fig. 3.
• The oligonucleotides occur naturally and are
used as primers during DNA replication and
for various other purposes in the cell.
Synthetic oligonucleotides can be made by
chemical synthesis and are essential for many
lab techniques, e.g., DNA sequencing, PCR, in
situ hybridization, nucleic acid probe, nucleic
acid hybridization, gene therapy.
• The polymers containing >100
ribonucleotides or deoxyribonucleotides are
called RNA and DNA (nucleic acids),
respectively.
Nomenclature of nucleic acids
• Direction: By convention, single strand of
nucleic acid is always written with the 5’
end at the left and 3’ end at the right
i.e.in 5’ →3’ direction.
• Sugar: In the chemical nomenclature,
the carbon atoms of sugars are
designated by primed numbers i.e.C-1’,
C-2’, C-3’ etc. to avoid confusion with the
base numbering system.
• Short hand notation: In short hand
notation of nucleotides, phosphate group
is symbolized by P, deoxyribose a vertical
line from C1’ at top to C5’ at bottom. The
connecting lines between nucleotides,
which pass through P, are drawn
diagonally from the middle (C3’) of
deoxyribose of one nucleotide to bottom
(C5’) of next (Fig. 4).
• The nucleoside and nucleotide
derivatives of deoxyribose are
distinguished by prefix ‘d’. Where clarity
is especially important, ribonucleosides
and ribonucleotides can similarly be
identified with the prefix’r’, e.g. ATP =
rATP.
Structural levels of nucleic acids
(a) Primary structure
• The nature, properties and function of the two
nucleic acids (DNA and RNA) depend on the
exact order of the purine and pyrimidine
bases in the molecule. This sequence of
specific bases is termed as the primary
structure. Thus, primary structure of nucleic
acid is its covalent structure and nucleotide
sequence.
(b) Secondary structure
• Secondary structure is the set of interactions
between bases, i.e., parts of which is strands
are bound to each other. In DNA double helix,
the two strands of DNA are held together by
hydrogen bonds. The nucleotides on one
strand base pairs with the nucleotide on the
other strand. The secondary structure is
responsible for the shape that the nucleic acid
assumes.
• Secondary structures in RNA, which exist
primarily in single stranded form, generally
reflect intra-molecular base interactions. Thus,
the secondary structures arise due to
following interactions:
• Complementary base pairing: It involves
stable and specific configurations of H-bonds
between bases in DNA. It is the predominant
force causing nucleic acid strands to associate.
The molecular basis of Chargaff’s rule is
complementary base pairing between A-T and
between G-C in double stranded DNA.
• (C) Tertiary structure
• Tertiary structures of nucleic acid reflect
interactions, which contribute to overall 3D
shape. Tertiary structure is the locations of the
atoms in three-dimensional space, taking into
consideration geometrical and steric
constraints. A higher order than the secondary
structure in which large-scale folding in a linear
polymer occurs and the entire chain is folded
into a specific 3-dimensional shape. There are 4
areas in which the structural forms of DNA can
differ.
•
•
•
•
Handedness - right or left
Length of the helix turn
Number of base pairs per turn
Difference in size between the major and
minor grooves.
• The tertiary arrangement of DNA's
double helix in space includes B-DNA, ADNA and Z-DNA.
(D) Quaternary structure
• The quaternary structure of nucleic acids is similar to
that of protein quaternary structure. Although some
of the concepts are not exactly the same, the
quaternary structure refers to a higher-level of
organization of nucleic acids. Moreover, it refers to
interactions of the nucleic acids with other molecules.
The most commonly seen form of higher-level
organization of nucleic acids is seen in the form of
chromatin which leads to its interactions with the
small proteins histones. Also, the quaternary structure
refers to the interactions between separate RNA units
in the ribosome or spliceosome.
DNA to Chromatin
• Deoxyribonucleic acid (DNA) is the genetic
material in all organisms, except few viruses
where RNA acts as the genetic material e.g.
retroviruses.
• In prokaryotic cells, DNA occurs in the
cytoplasm and is the only component of the
chromosome. In eukaryotic cells, DNA is
largely confined to the nucleus and is the main
component in chromosome. It is combined
with simple proteins to form deoxy-ribonucleoproteins (DNP).
• A small quantity of DNA also occurs in
some cytoplasmic organelle such as
mitochondria and chloroplast. This extranuclear DNA is naked as in prokaryotic
DNA. The DNA content is fairly constant
in all the cells of a given species. Just
before cell division, however, the amount
of DNA is doubled. The gametes have
half the amount of DNA as they contain
half the number of chromosomes.
• The amount of DNA per nucleus is
constant in all body cells of a given
species. Mirsky and Vendrely estimated
that there is some 6x10-9 mg of DNA per
nucleus in diploid somatic cells of
mammals and 3 x 10-9 mg of DNA per
nucleus in haploid gametes (eggs and
sperms).
EVIDENCES THAT DNA IS GENETIC
MATERIAL
• DNA was first extracted from nuclei in
1870 named ‘nuclein’ after their source.
Chemical analysis determined that DNA
was a weak acid rich in phosphorous. Its
name provides a lot of information about
DNA: deoxyribose nucleic acid: it
contains a sugar moiety (deoxyribose), it
is weakly acidic, and is found in the
nucleus.
• Because of its: nuclear localization subsequent
identification as a component of chromosomes it was
implicated as a carrier of genetic information.
Bacterial transformation implicates DNA as the
substance of genes.
• 1928 Frederick Griffith experiments with smooth (S),
virulent strain Streptococcus pneumoniae, and rough
(R), non-virulent strain.
• Bacterial transformation demonstrates transfer of
genetic material.
• In 1944 – Oswald Avery, Colin MacLeod, and MacIyn
McCarty determined that DNA is the transformation
Material.
Griffith experiment: transformation of bacteria
Griffith’s Experiment
• Griffith observed that live S bacteria could kill mice
injected with them.When he heat killed the S variants
and mixed them with live R variants, and then injected
the mixture in the mice, they died. Griffith was able to
isolate the bacteria from the dead mice, and found
them to be of the S variety. Thus the bacteria had
been Transformed from the rough to the smooth
version. The ability of a substance to change the
genetic characteristics of an organism is known as
transformation. Scientists set out to isolate this
‘transforming principle’ since they were convinced it
was the carrier of the genetic information.
Avery, MacLeod, McCarty Experiment: Identity of the
Transforming Principle
• Hershey and Chase experiment
• In 1952 Alfred Hershey and Martha Chase
provide convincing evidence that DNA is
genetic material.
• Waring blender experiment using T2
bacteriophage and bacteria.
• Radioactive labels 32P for DNA and 35S for
protein.
Hershey and Chase experiment
• Performed in 1952, using bacteriophage, a type of virus
that have a very simple structure: an outer core and an
inner component. The phage is made up of equal parts
of protein and DNA. It was known that the phage infect
by anchoring the outer shell to the cell surface and then
deposit the inner components to the cell, infecting it.
• Scientists were interested in finding out whether it was
the protein component or the DNA component that got
deposited inside the infected cell. By incorporating
radiolabel either in the protein or the DNA of the
infecting phage, they determined that the DNA was
indeed introduced into the infected bacteria, causing
proliferation of new phage.
Chromosomes
• The cell nucleus contains the majority of the
cell's genetic material, in the form of multiple
linear DNA molecules organized into structures
called chromosomes (complex very compact
structures of DNA in association with various
simple proteins).
• During most of the cell cycle, DNAs are organized
in a DNA-protein complex known as chromatin,
and during cell division the chromatin can be
seen to form the well defined chromosomes
familiar from a karyotype.
• A small fraction of the cell's genes are located
instead in the mitochondria.
• A chromosome is an organized structure of
DNA and protein that is found in cells. It is a
single piece of coiled DNA containing many
genes, regulatory elements and other
nucleotide sequences.
• In eukaryotes, nuclear chromosomes are
packaged by proteins into a condensed
structure called chromatin. This allows the
very long DNA molecules to fit into the cell
nucleus.
• Chromosomes may exist as either duplicated
or unduplicated. Unduplicated chromosomes
are single linear strands, whereas duplicated
chromosomes (copied during synthesis phase)
contain two copies joined by a centromere.
Compaction of the duplicated chromosomes
during mitosis and meiosis results in the
classic four-arm structure.
Chromatin Structure
• Chromatin is the chromosomal material
found in nuclei of cells of eukaryotic
organisms. Each chromatin consists of a
single double-stranded DNA in
combination with a nearly equal amount
of small basic proteins called histones
and smaller amounts of non-histone
proteins (most of which are acidic and
larger in size than histones) and a small
quantity of RNA molecules.
• The double stranded helix of DNA has a
length that is thousands of times larger
than the diameter of the cell nucleus.
Histones facilitate condensation
(compactness) of DNA, so that all
chromosomes can fit into the nucleus.
The chromatin is composed of repeating
units of dense spherical particles called
nucleosomes (about 10 nm in diameter)
connected by DNA filaments.
• There are two types of chromatin.
Euchromatin is the less compact DNA form,
and contains genes that are frequently
expressed by the cell. The other type,
heterochromatin, is the more compact form,
and contains DNA that are infrequently
transcribed.
• Histones organize DNA, condense it and
prepare it for further condensation by
nonhistone proteins. This compaction is
necessary to fit large amounts of DNA
(2m/6.5ft in humans) into the nucleus of a
cell.
• Both histones and nonhistones are
involved in physical structure of the
chromosome. Histones are abundant,
small proteins with a net (+) charge. The
five main types are H1, H2A, H2B, H3, and
H4. By weight, chromosomes have equal
amounts of DNA and histones. Histones
are highly conserved between species H1
less than the others.
• A possible nucleosome structure
• Chromatin formation involves histones and
DNA condensation so it will fit into the cell,
making a 10-nm fiber. Chromatin formation
has two components: two molecules each of
histones H2A, H2B, H3, and H4 associate to
form a nucleosome core and DNA wraps
around it 1-3 or 4 times for a 7-fold
condensation factor.
• Nucleosomes form the fundamental
repeating units of eukaryotic chromatin
which is used to pack the large eukaryotic
genomes into the nucleus. In mammalian
cells approximately 2 m of linear DNA have
to be packed into a nucleus of roughly 10 µm
diameter. Nucleosomes are folded through a
series of successively higher order structures
to eventually form a chromosome. These
compacts DNA and creates a regulatory
control which ensures correct gene
expression.
• Fully condensed chromosome is 10.000fold shorter and 400-fold thicker than
DNA alone.
The many different orders of chromatin packing that give rise to the highly
condensed metaphase chromosome:
Centromeric and Telomeric DNA
• These are eukaryotic chromosomal
regions with special functions.
Centromeres are the site of the
kinetochore, where spindle fibers attach
during mitosis and meiosis. They are
required for accurate segregation of
chromatids.
• Kinetochores are regions found in
chromosomes. They contain highly
repetitive DNA sequences, and are
bound to by many proteins. During cell
division, microtubules are attached to
these regions for chromosome
segregation (kinetochore). Kinetochores
are equivalent to the primary
constriction sites of chromosomes in
higher eukaryote.
• Telomeres are located at the ends of
chromosomes, and needed for
chromosomal replication and stability.
Generally composed of heterochromatin,
they interact with both the nuclear
envelope and each other. All telomeres in
a species have the same sequence.
Centromeres and telomeres are both
Heterochromatin.
Anatomy of Chromosome
• Diagram of a duplicated and condensed
metaphase eukaryotic chromosome.
1. Chromatid, one of the two identical
parts of the chromosome after S-Phase.
2. Centromere, the point where the two
chromatids touch, and where the
microtubules attach.
3. Short arm (is known as p).
4. Long arm (is known as q).
Functions of DNA
• DNA is the very basis of life and has five-fold roles:
• It carries hereditary characters from parents to offspring.
• It enables the cell to maintain grow and divide by directing
the synthesis of structural proteins.
• It controls metabolism in the cell by directing the formation of
necessary enzymatic proteins.
• It contributes to the evolution of the organism by undergoing
gene mutations (changes in the sequence of base pairs).
• It brings about differentiation of cells during development.
Only certain genes remain functional in particular cell. This
enables the cells having similar genes to assume different
structure and function.
Human Genome
• Human genome is the totality of DNA
characteristic of all the 23 pairs of
chromosomes.
• The human genome has about 3x109 bps
in length.
• 97% of the human genome is non-coding
regions called introns. 3% is responsible
for controlling the human genetic
behavior. The coding region is called
extron.
• There are totally about 40.000 genes,
over 5.000 have been identified. There
are much more left.
• Human Genome Project is to identify the
DNA sequence (every bp) of human
genome (only a few individuals). For
human being, most of the place in
human genome are the same. Only a
very small part is different among
different individuals.
• Genes or DNA sequences them self are
not control the phenotypes, they control
the phenotypes through protein. Protein:
like the DNA molecule that is a chain of
base pair, each protein molecule is a
linear chain of subunits called amino
acids.
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