DMD1001

CELL AND
MOLECUL
AR
BIOLOGY

Dr Kereann Nelson
Important Information
Recommended text
• Molecular Biology of the Cell- Bruce Alberts, Alexander D. Johnson, et al.

Test 1- Week 7
Test 2- Week 12

Presentations- Week 12
 Unit 1

The Structure of DNA

DNA is a nucleic acid

• So is RNA

• Nucleic acids first discovered in 1868 by

Friedrich Meischer
• Nuclei of white blood cells
• He called this substance “nuclein”

• Overall small negative charge

• Mild acid
 1. Stores genetic information as a linear
sequence of bases.
ATGGTTCGG…..

2. Replicates itself during cell division to

Functions of provide a copy of the genome to each
daughter cell.
DNA
3. Provides a template for gene
expression (RNA and protein
synthesis).
• Let’s define the word “template”….
 Basic DNA
Structure

A polymer of deoxyribonucleotides:
Let’s define “polymer”….

• Nucleic Acids are named

according to the sugars
present.

• Sugars:
1. deoxyribose (DNA)
2. ribose (RNA).
 DNA
Components:
• Long molecule made up of many repeating units.
• These units are called nucleotides

• Nucleotides are made up of 3 components

covalently bonded together:
1. A Sugar
2. A phosphate group
3. A nitrogenous base

• [Nucleoside is a base + a sugar]

• Let’s define a covalent bond…

 Purines Pyrimidines
• Double ring bases. • Single ring bases.
Bases:

Nitrogen-
containing
compounds

Images: Blausen.com staff (2014). "Medical

gallery of Blausen Medical 2014". WikiJournal of
Medicine 1 (2). DOI:10.15347/wjm/2014.010.
ISSN 2002-4436.
 • In RNA Uracil
replaces
Thymine

Basic DNA Structure

 • After the primary bases have been incorporated into
The bases the nucleotide they can get chemically modified for
different reasons in different ways.
previously • Modified bases include:
• 5-methylcytosine (m5C)- Very common in DNA
discussed • We will discuss its significance later in the
course
are • In RNA some modified bases are:
• pseudouridine (Ψ), dihydrouridine (D), inosi
Primary ne (I), and 7-methylguanosine (m7G)

Bases • Modification of the primary bases due to mutations

can affect base-pairing properties.
Nucleotide Formation
 1. Adenine + Pentose Sugar = Adenosine
(Adenine Nucleoside)

Formation 2. Adenosine + Phosphate group = Adenylic Acid

of or Adenosine Monophosphate (AMP).

Nucleotides (Adenine Nucleotide)

- Using ATP 3. Adenosine Monophosphate + Phosphate =

as an Adenosine Diphosphate (ADP).

example 4. Adenosine Diphosphate + Phosphate =

Adenosine Triphosphate (ATP).
 Nucleosides vs Nucleotides
Base Nucleoside Deoxynucleoside Deoxynucleotide
Adenine Adenosine 2-deoxyadenosine 2-deoxyadenosine triphosphate
(dATP) ATP if RNA
Cytosine Cytidine 2-deoxycytidine 2-deoxycytidine triphosphate
(dCTP) CTP if RNA
Guanine Guanosine 2-deoxyguanosine 2-deoxyguanosine triphosphate
(dGTP) GTP if RNA
Thymine Thymidine 2-deoxythymidine 2-deoxythymidine triphosphate
(dTTP) TTP if RNA
Uracil Uridine Not usually found in DNA Not usually found in DNA
RNA- uridine triphosphate
UTP
Composition of DNA

source:
http://nptel.ac.in/courses/104103018/module1/lec6/5.html
 • Each deoxynucleoside
triphosphate (dNTP) is
linked to the next by
phosphodiester
bonds.

Phosphate group and

phosphodiester bond
 The
phosphodiester
bond

• A condensation reaction
• Esterification.
• Bond formed between the free
OH at Carbon 3 (3’) and the
alpha phosphate at Carbon 5
(5’) of the dNTP.
• Links 3’ carbon of one
sugar with 5’ carbon of
another.

• Results in 5’ and 3’ ends.

 Chargaff’s
Rules

• The C&G : A&T ratio varies

among species but the ratio
of adenine to thymine is
always equal, as is the ratio
of cytosine to guanine.
• A=T
• C=G
• but AT≠GC.
• %GC varies across
genomes.
• Mycobacterium have
GC ~ 73%
DNA is double-
stranded
• Double helix with strands running antiparrallel.
• Sugar-phosphate chains on the outside (sugar-
phosphate backbone).
• Core is formed from bases held together by H-
bonds.
• Putting the hydrophobic bases inside protects
them from interaction with water or certain
hydrophilic compounds.

• Definitions?
 1. The Hydrogen bonds between the bases
(previously discussed).
2. Hydrostatic forces:
• The sugar-phosphate backbone of DNA is polar, and
therefore hydrophilic.
DNA molecule
• The bases, are relatively non-polar and therefore
is a stable hydrophobic.
double strand • These hydrostatic forces have a very stabilising effect
due to: on the overall structure of the DNA double helix
• Therefore there is a strong pressure holding the two
strands of DNA together.

Definitions?
Complementary base-pairing
• 3 H-bonds G-C
• 2 H-bonds A-T

• Hydrogen bonds are not chemical bonds

• can be easily disrupted
• This permits the DNA strands to separate for
transcription (copying DNA to RNA) and replication
(copying DNA to DNA).

• DNA with higher G-C content is more stable.

• i.e. strands (or that region of DNA) harder to separate.
  The helix makes a turn every 3.4 nm
 The distance between two neighboring base
pairs is 0.34 nm.
The DNA  Hence, there are about 10 pairs per turn.
Double  The intertwined strands make two grooves of
different widths:
Helix  The major groove

 The minor groove

 source:
http://bioserv.fiu.edu/~walterm/GenBio2004/new_chap14_DNA/dna_and_heredity.htm
DNA exists in different shapes (conformations)

Conformation
depends on:
1. DNA sequence.
A-DNA, B-DNA, and Z-DNA are 2. Amount and direction of
the forms observed in cells. supercoiling.
Under physiological 3. Chemical modifications of the
bases.
conditions, most DNA
DNA exists in other possible is in the B form. 4. Solution conditions (e.g.
conformations. concentration of metal ions,
• E.g. D-DNA salt concentration and level of
hydration).
 B-DNA

• First described by James D.

Watson and Francis Crick in
1953.
• Watson and Crick based
their model (B-DNA helix)
on x-ray diffraction patterns
of DNA fibres.
 Watson and
Crick’s Model of
DNA

• Rosalind Franklin and

Maurice Wilkins
 Exposed DNA crystals to X-
rays.
 The pattern generated
when the X-rays bounce
off different atoms gives
insight into the structure
of the molecule.
 1. Rosalind Franklin
discovered during
crystal formation
that DNA exists as A-
DNA and B-DNA. She
experimented until
she separated the
two conformations.
2. Ladder-like pattern in
crystallography of
what would later be
identified as
nucleotides.
What information did X- 3. “X” shape indicated
ray crystallography that DNA had a
helical shape.
provide?
 4. “X” pattern was
consistent
indicating that DNA
is uniform in
diameter.
5. Dimensions of DNA
and number of
bases per turn.
6. Orientation of
bases and sugar-
What information did X- phosphate
backbone.
ray crystallography
provide (cont.)?
 In 1953 James Watson and Francis Crick

• Saw Franklin and Wilkin's picture of the X-ray and made

an accurate model - a double helix with rungs
connecting the two strands

Watson How were the bases bonded together?

and Crick’s What is the best orientation of the bases to

arrive at the shape seen in the X-ray?

Model of •
•
Chargaff’s Rule applied.
Adenine and Guanine were purines (2 rings).
DNA •
•
Thymine and Cytosine were pyrimidines (1 ring).
If purines were linked together and pyrimidines
were linked together, the shape would not be
uniform.
• If A-T and G-C DNA has a uniform shape.
• Chargaff’s rules show A=T, G=C.
Remembering
Rosalind
• Franklin’s work provided the basis for the model
created by Watson and Crick.
• The model generated by Watson and Crick also
used information from Avery, Chargaff, Griffiths,
and others.
• The Nobel Prize was awarded to Watson, Crick, and
Maurice Wilkins.
• Rosalind Franklin did not receive the prize because
she had died of cancer in 1958.
• Maurice Wilkins shared the prize with Watson and
Crick because of his work with Franklin. Her
accomplishment should never be forgotten.
B-DNA
 Most common form of DNA under the
conditions found in cells.

Has a major and minor groove

Summary
of B-form
of DNA Has 10 base pairs per turn

One turn spans 3.4 nm

 • The results of x-ray diffraction studies of dehydrated
DNA fibres revealed a different form called A-DNA.

• Appears when the amount of water in the solution is

reduced to less than 75%.
• Results, for example, at higher salt
concentrations or when alcohol is added.
A-DNA 1. Also a right-handed double helix made up of
antiparallel strands held together by Watson-Crick
base-pairing.
2. Helix is wider and shorter than the B helix.
3. Base pairs are tilted rather than perpendicular to
the helix axis.
A-DNA (cont.)

• A shallow, wider
minor groove.
• A narrower, deeper
major groove.
• Has 11 bases per
turn.
• One turns spans 2.3
nm.
 • The A-DNA helix is not confined to dehydrated
A-DNA in DNA.
• In the cell it can be found in:
the cellular 1. Double-stranded regions of RNA.
environme 2. Some RNA-DNA hybrids adopt a double-
nt helical form very similar to that of A-DNA.
 • Alexander Rich and his associates discovered a
third type of DNA helix when they solved the
structure of dCGCGCG.
• They found that this hexanucleotide forms a
duplex of antiparallel strands held together by
Watson-Crick base-pairing, as expected.
Z-DNA • What was surprising, however, was that this
double helix was left-handed, in contrast with
the right-handed screw sense of the A and B
helices.
• Furthermore, the phosphates in the backbone
zigzagged; hence, they called this new form Z-
DNA.
 • Adopted by short oligonucleotides that have
sequences of alternating pyrimidines and purines.
• Structure is maintained by high salt concentrations.
• High salt concentrations minimize the electrostatic
repulsion between the backbone phosphates,
which are now closer to each other than they
would be in A- and B-DNA.
Z-DNA • This structure is therefore best observed if there are
alternating G-C sequences in alcohol or high salt
solution.
• Biological role-
• Regulation of gene expression
• Certain aspects of transcription
• Increased amounts linked to Alzheimer disease.
 1. Bases seem to zigzag.
2. The strands turn about the helical axis in a
left-handed spiral.
Z-DNA 3. Has a narrow deep groove
4. Has 12 bases per turn.
5. One turn spans 4.56 nm
Chromosomes:
 • Chromosomes – highly coiled condensed
packages of DNA
• Present in both eukaryotes and prokaryotes
Chromoso • Eukaryotes – linear
mes • Prokaryotes – most closed circular
• The human genome has 3,000,000,000 base
pairs packed into 23 chromosomes.
 • DNA is packed into chromosomes with the help
of proteins - histones.

Organizatio • The histones associate with DNA forming

n of structures called nucleosomes.

Eukaryotic • Each nucleosome complex consists of a beadlike

Genomes structure with 146 base pairs of DNA wrapped
around a disc-shaped core of eight histone
molecules.
Nucleosome
 • Nucleosomes are ~11nm in diameter.

• The packed nucleosome state occurs when a

ninth histone called H1, associates with the
linker DNA packing adjacent nucleosomes
Chromoso together forming a 30nm diameter thread.

mes
• In the extended chromosome , these 30nm
diameter threads form large coiled loops held
together by a set of non-histone scaffolding
proteins.
 Eukaryotic
Genomes

• Organised into
multiple
chromosomes -
varying in size and
numbers depending
on the species:
 • The number, size and shape of the
chromosomes of a somatic cell arranged in a
standard manner.
• The normal human karyotype has 46
chromosomes.
KARYOTYPE • Position of centromere - arm length ratio.
• Chromosomes are matched with their
homologue.
• Secondary constrictions (nucleolar organisers)
and satellites.
 • Short arm is labeled P (French for petit)
• Long arm is labeled Q

• Human chromosomes are divided into 7 groups +

Chromosomes sex chromosomes:
are always A. 1-3 Large metacentric 1,2 or submetacentric
arranged with B. 4,5 Large submetacentric, all similar
the short arm on C. 6-12, X Medium sized, submetacentric - difficult
top in D. 13-15 medium-sized acrocentric plus satellites
karyotypes E. 16-18 short metacentric 16 or submetacentric
17,18
F. 19-20 Short metacentrics
G. 21,22,Y Short acrocentrics with satellites. Y no
satellites.
 • Metacentric:
• centromere is median or near median
• chromosome has two well defined arms with a length ratio varying
from 1:1 to 2.5:1
• Acrocentric:
• centromere is close to one end of the chromosome

Classifica • one arm is substantially smaller than the other and the arm ratio
ranges from 3:1 to 10:1

tion of • Telocentric:
• centromere is a strictly terminal entity and the chromosome is one
chromoso armed

mes
 • Most have covalently closed, circular
chromosomes and plasmids
Bacterial • Not all bacteria have a single circular
chromosome:
Genome • Some bacteria have multiple circular
Organizatio chromosomes

n • Many bacteria have linear chromosomes and

linear plasmids.
 • The genetic material of a bacterium lies in the
cytoplasm and is not surrounded by a nuclear
envelope.
• Nucleoid

Bacterial • In most species it is contained in a single circular

DNA molecule.
Genome
• If stretched out to its full length, this molecule
would be 1000 times longer than the cell itself.

• Unlike eukaryotic chromosomes, the bacterial DNA

has only a small amount of proteins associated
with it.
 • In addition to the genomic DNA, most
bacteria have a small amount of genetic
information present as one or more
plasmids – extrachromosomal circular DNA.

• Plasmids can replicate independently of the

genomic DNA or become integrated into it.

• Bacterial plasmids frequently have genes

that code for:
• Catabolic enzymes.
• Genetic exchange.
• Resistance to antibiotics.

Plasmids
 Viral Genomes
Virus genomes may contain their genetic information encoded in either DNA or RNA
 • Many of the DNA viruses of eukaryotes closely
resemble their host cells in terms of the biology
of their genomes:
Viral • Some DNA virus genomes are complexed
with cellular histones to form a chromatin-
Genomes like structure inside the virus particle.
 Genome may be :
• Circular or linear
• Segmented – (two or more separate molecules of
nucleic acid)
• Single-stranded
• Double-stranded
• Double-stranded with regions of single-strandedness
Viral • DNA viruses

Genomes • RNA viruses

• Positive sense (acts like mRNA and can be immediately
make proteins)
• Negative sense (needs transcription to a complementary
mRNA)
• Ambisense (has regions of + and – polarity)
• Both DNA and RNA (at different stages in the life
cycle)
• Reverse transcribing viruses (RNA DNA RNA
Protein)
 1. dsDNA viruses
• (e.g. Herpesviruses)
2. ssDNA viruses (+)sense DNA
• (e.g. Parvoviruses)
3. dsRNA viruses
• (e.g. Reoviruses - Rotavirus)
4. (+)ssRNA viruses
• (+)sense RNA (e.g. Picornaviruses -Enterovirus)
Viral Genomes 5. (-)ssRNA viruses
• (-)sense RNA (e.g. Orthomyxoviruses – Influenza A,
B, C)
6. ssRNA-RT viruses
• (+)sense RNA with DNA intermediate in life-cycle
(e.g. Retroviruses – HIV, leukaemia and tumour
viruses)
7. dsDNA-RT viruses
• (e.g. Hepadnaviruses – hepatitis B)
 Unit of inheritance.

Genes determine traits.

Genes
A sequence of nucleotides which provide a
cell with the instructions for the synthesis
of a specific protein or a type of RNA.

Most are 1,000 to 4,000 nucleotides long

(may be shorter or significantly longer).
Genes
• Most eukaryotic
genes contain
non-coding
regions - introns
 • Genome sizes are typically expressed as follows:
• Kilobase (kb)
• 103 nucleotide pairs (double-stranded)
or
• 103 nucleotides (single-stranded)

• Megabase (Mb)
Genome size • 106 nucleotide pairs (double-stranded)
or
• 106 nucleotides (single-stranded)
• Gigabase (Gb)
• 109 nucleotide pairs (double-stranded)
or
• 109 nucleotides (single-stranded)
 Genome • The genome size of an organism may also be
measured as its “C-value”.
Size and • the total amount of DNA contained within a
single (i.e., haploid) set of its chromosomes.
Evolutionar • The C-value measured chemically in picograms (pg)
y of DNA or by DNA reassociation, when it is generally
given in base pairs or daltons.
complexity: • 1 pg = 0.965 x 109 bp = 6.1 x 1011 daltons.

The C- • There is enormous variation in the range of C

value values, from as little as a mere 104 bp for
mycoplasma to as much as 1011bp for some plants
Paradox and amphibians.
 • Viral genomes are typically in the range 100 –
1000 kb
• Bacterial genomes are typically in the range 1 –
Typical 10 Mb
Genome • Eukaryotic genomes are typically in the range
100 -1000 Mb
Size • (smallest eukaryotic genomes are about 10
Mb)
 Genome
size (cont.)
 Molecular Biology
in Dentistry

• Teeth are amongst the hardest

structures of the human body.
• Resistant to adverse
conditions such as
incineration, immersion,
trauma, mutilation,
decomposition and hence,
used in forensic
investigations.
• They are a valuable source of
DNA (pulp) as other parts of
the body may get destroyed or
degraded.
 • DNA profiling and forensic dentistry.
• The fast technological advancements in DNA
Molecular research have revolutionized the field of
forensic medicine and the present work was
Biology in undertaken to provide an insight in to the
Dentistry recent concepts of DNA profiling in Forensic
dentistry.
 • There are more than 600 different types of
bacteria present in the human mouth at any
given time.
• 13 strains are the most common pathogenic
Molecular bacteria which cause gum disease.
Biology in • Periodontal pathogens are opportunistic.
• Some of the catalysts may include emotional or
Dentistry physical stress, hormonal imbalances, smoking,
acute infections and chronic medical conditions
such as asthma and diabetes.
 • Until recently periodontal disease was treated
by identifying clinical signs and symptoms.
• It can take years for a patient to perceive a
problem.
Molecular • Now Polymerase Chain Reaction uses molecular
biology technology to provide a solid method of
Biology in identifying the disease.
Dentistry • Now able to specifically identify the oral
bacteria that can cause infections before they
become an issue.
• Able to prescribe treatment before the
symptoms become widespread.
 • Gum disease is also believed to be a precursor
to other health problems: heart disease and
stroke.
• Molecular studies of pathogens implicated in
Molecular periodontal issues.
Biology in • Also research aimed at generating human
periodontal ligament stem cells for treatment
Dentistry of periodontal disease.
• Studies of oral cancers.
• Genetics of craniofacial and dental anomalies.
Molecular Biology in Dentistry
Salivary Diagnostics
• Saliva is said to be a “mirror of the body”
because it is an indicator of health not just in the
oral cavity but throughout the body.
• The molecular composition of saliva includes
therapeutic, hormonal, immunologic, and
toxicological molecules, which can provide vital
clues to systemic health.
• Saliva has been used informally as a clinical
diagnostic medium for more than 2,000 years.
 • Historically, the viscosity and odor of saliva
Molecular were used as diagnostic symptoms for certain
diseases.
Biology in • In diagnostics, saliva is an excellent alternative
to serum or urine since it contains sufficient
Dentistry quantities of disease biomarkers, ribonucleic
Salivary acid (RNA), and deoxyribonucleic acid (DNA),
and the collection method is non-invasive, safe,
Diagnostics and easy.
 • Saliva-based diagnostic technologies have the potential
to improve dental care by detecting oral infections
early. The ultimate demineralization of teeth seen in
dental caries is caused by acids produced by bacteria,
particularly Streptococcus mutans and Lactobacilli.
• Prediction of caries risk is often difficult because of the
complex interactions of the host and its environment,
Salivary as well as behavioral risk factors. Changes in buffer
capacity, salivary composition, rate of salivary flow, and
Diagnostics pH are all risk factors for caries.
• One of the most significant predictors of caries is the
relative level of S. mutans and Lactobacilli colonization,
which can be determined through saliva or plaque
samples.
Two DNA-based saliva tests are
available from OralDNA® Labs:

• MyPerioPath® and MyPerioID® PST®.

• My PerioPath uses a saliva sample to identify
the type and concentration of the specific
bacteria that cause periodontal diseases.
• MyPerioID PST test also uses saliva to
determine a patient’s genetic susceptibility to
periodontal diseases and which patients are at
higher risk of more serious periodontal
infections.
 SAMPLE • Sample-Oral DNA report
ORAL DNA • www.oraldna.com/pdf/OralDNA
Labs-MyPerioPath-report.pdf
REPORT
 • http://www.ncbi.nlm.nih.gov/books/NBK22585
/
Required
Reading • http://www.ncbi.nlm.nih.gov/pmc/articles/PMC
3125955/