NUCLEIC ACIDS

DNA RNA

DEOXYRIBO RIBO
NUCLEIC NUCLEIC
ACID ACID
 DNA
• Discovery of the DNA double helix

A. 1950’s
B. Rosalind Franklin - X-ray photo of DNA.
C. Watson and Crick - described the
DNA molecule from Franklin’s X-ray.

1962 Nobel Prize

 Question:
• What is DNA?
 Deoxyribonucleic Acid (DNA)
• Made up of nucleotides in a DNA double helix.
• Nucleotide:
1. Phosphate group
2. 5-carbon sugar = deoxyribose
3. Nitrogenous base (4 different ones)
• ~2.4 nm wide

• ~1.8 meters long (in one cell) (~6 feet)

 DNA Nucleotide
Phosphate
Group

O 5’ Base
O=P-O CH2
O
O
N
Nitrogenous base
C4 Sugar C
1
(A, G, C, or T)
Sugar
(deoxyribose)
C3’ C2
 DNA Nucleoside

5’ Base
HOCH2

O
N
Nitrogenous base
C4’ Sugar C1’ (A, G, C, or T)
Sugar
(deoxyribose)
C3’ C2’
DNA Double Helix

“Steps of ladder”

Nitrogenous
Base (A,T,G or C)

“Legs of ladder”

Phosphate &
Sugar Backbone
 DNA Double Helix
5 O 3

3 O
P 5 P
5
O
1 G C 3
2
4 4
2 1
3 5
O
P P
5
T A 3

O
5
P 3 P
 Nitrogenous Bases

• PURINES (double ring)

1. Adenine (A)
A or G
2. Guanine (G)

• PYRIMIDINES (one ring)

3. Thymine (T)
4. Cytosine (C) T or C
COMPLEMENTARY BASE-PAIRING
Base # of
Purines Pyrimidines Pairs H-Bonds
Adenine (A) Thymine (T) A=T 2

Guanine (G) Cytosine (C) C G 3

3 H-bonds

G C
COMPLEMENTARY BASE-PAIRING
3 H-bonds

G C

2 H-bonds

T A
 Chargaff’s Rule
• Adenine must pair with Thymine

T A

• Guanine must pair with Cytosine

G C
 Non-dividing DNA PACKING Dividing
loosely coiled tightly coiled
HISTONES

Organization
of DNA in
Chromosomes
NUCLEOSOME
 CHROMATIN PACKING
DNA
in
nucleus
2 meters
~ 6 feet

50,000
times
shorter
 Question:
• How does RNA (ribonucleic acid) differ
from DNA (deoxyribonucleic acid)?
acid)
 RNA differs from DNA

1. RNA has a sugar ribose

DNA has a sugar deoxyribose

2. RNA contains uracil (U) (pyrimidine)

DNA has thymine (T) (pyrimidine)

3. RNA molecule is single-stranded

DNA is double-stranded
 Types of RNA
• Three major types of RNA:
A. messenger RNA (mRNA)
B. transfer RNA (tRNA)
C. ribosome RNA (rRNA)

• Remember: all produced in the nucleus!

DNA function
genetic code

RNA function
protein synthesis
 Protein Synthesis
Instructions for making specific proteins is found in the
DNA (our genes)
DNA PROTEIN

HUMAN CHROMOSOMES
23 PAIRS
 Protein Synthesis
• The production (synthesis) of proteins.
proteins
• 3 phases:
phases
1. Transcription
2. RNA processing
3. Translation
• Remember: DNA → RNA → Protein
DNA → RNA → Protein
Nuclear
DNA membrane

Transcription
Pre-mRNA

RNA Processing
mRNA

Ribosome

Translation

Protein
 1. Transcription
• The transfer of information in the nucleus from a
DNA molecule to an RNA molecule.
• Only 1 DNA strand serves as the template
• Starts at promoter DNA (TATA box)
• Ends at terminator DNA (stop)
• When complete, pre-RNA molecule is released.
 Question:
• What is the enzyme responsible
for the production of the RNA
molecule?
 Answer: RNA Polymerase
• Separates the DNA molecule by breaking the
H-bonds between the bases.
• Then moves along one of the DNA strands and
links RNA nucleotides together.
 1. Transcription
DNA

RNA Polymerase

pre-mRNA
 1. TRANSCRIPTION

RNA synthesis

http://www.schenectady.k12.ny.us/putman/biology/data/transcription/complete.html
 Question:
• What would be the complementary
RNA strand for the following DNA
sequence?

• DNA 3’- GCGTATG - 5’

 Answer:

• DNA 3’- GCGTATG - 5’

• RNA 5’- CGCAUAC - 3’

2. RNA Processing
Nuclear
DNA membrane

Transcription
Pre-mRNA

RNA Processing
mRNA

Ribosome

Translation

Protein
 2. RNA Processing
• Maturation of pre-RNA molecules.
• Also occurs in the nucleus.
• Introns spliced out by spliceosome enzyme
and exons come together.
• End product is a mature RNA molecule that
leaves the nucleus to the cytoplasm.
 2. RNA Processing
pre-RNA molecule

exon intro exon intron exon

n

intron intron

exon exon exon

spliceosome spliceosome

exon exon exon

Mature RNA molecule

Pre-mRNA Processing (Splicing)
Human Genome Sequence
unexpected "Small" Size
~25,000 genes
Less than 1.5% of human genome encodes proteins
non protein coding sequences
 Types of RNA
• Three major types of RNA:
A. messenger RNA (mRNA)
B. transfer RNA (tRNA)
C. ribosome RNA (rRNA)

• Remember: all produced in the nucleus!

 A. Messenger RNA (mRNA)
• Carries the information for a specific protein.
protein
• Made up of 500 to 1000 nucleotides long.
• Made up of codons (sequence of three bases:
AUG - methionine).
• Each codon,
codon is specific for an amino acid.
acid
 A. Messenger RNA (mRNA)
start
codon

mRNA A U G G G C U C C A U C G G C G C A U A A

codon 1 codon 2 codon 3 codon 4 codon 5 codon 6 codon 7

protein methionine glycine serine isoleucine glycine alanine stop

codon

Primary structure of a protein

aa1 aa2 aa3 aa4 aa5 aa6

peptide bonds
GENETIC CODE: CODONS

Third letter
 B. Transfer RNA (tRNA)
• Made up of 75 to 80 nucleotides long.
• Picks up the appropriate amino acid floating in the
cytoplasm (amino acid activating enzyme)
enzyme

• Transports amino acids to the mRNA.

mRNA
• Have anticodons that are complementary to mRNA
codons.
codons
• Recognizes the appropriate codons on the mRNA
and bonds to them with H-bonds.
 B. Transfer RNA (tRNA)
amino acid
attachment site methionine amino acid

U A C

ANTI-CODON
 C. Ribosomal RNA (rRNA)
• Made up of rRNA is 100 to 3000 nucleotides
long.
• Important structural component of a ribosome.
• Associates with proteins to form ribosomes.

http://www.bio.miami.edu/dana/250/ribosomes.jpg
http://en.wikipedia.org/wiki/Ribosomal_RNA
 Ribosomes
• Large and small subunits.
• Composed of rRNA (60%) and proteins (40%).
• Both units come together and help bind the
mRNA and tRNA.
• Two sites for tRNA
a. P site (first and last tRNA will attach)
attach
b. A site
 Ribosomes

Large
subunit
P A
Site Site

mRNA
A U G C U A C U U C G

Small subunit
 3. Translation
Nuclear
DNA membrane

Transcription
Pre-mRNA

RNA Processing
mRNA

Ribosome

Translation

Protein
 3. Translation
• Synthesis of proteins in the cytoplasm

• Involves the following:

1. mRNA (codons)
2. tRNA (anticodons)
3. rRNA
4. ribosomes
5. amino acids
 3. Translation
• Three parts:
1. initiation:
initiation start codon (AUG)
2. elongation:
elongation
3. termination:
termination stop codon (UAG)

• Let’s make a PROTEIN!!!!.

PROTEIN!!!!
 3. Translation

Large
subunit
P A
Site Site

mRNA
A U G C U A C U U C G

Small subunit
 Initiation
aa2
aa1

2-tRNA
1-tRNA
G A U
anticodon U A C
hydrogen A U G C U A C U U C G A
bonds codon mRNA
 Elongation
peptide bond
aa3
aa1 aa2

3-tRNA

1-tRNA 2-tRNA G A A
anticodon U A C G A U
hydrogen A U G C U A C U U C G A
bonds codon mRNA
 aa1 peptide bond
aa3
aa2

1-tRNA

U A C 3-tRNA
(leaves)
2-tRNA G A A

G A U
A U G C U A C U U C G A
mRNA

Ribosomes move over one codon

 peptide bonds
aa1 aa4

aa2 aa3

4-tRNA

2-tRNA 3-tRNA G C U

G A U G A A
A U G C U A C U U C G A A C U
mRNA
 peptide bonds
aa1 aa4
aa2

aa3

2-tRNA
4-tRNA
G A U
(leaves) 3-tRNA G C U

G A A
A U G C U A C U U C G A A C U
mRNA

Ribosomes move over one codon

 peptide bonds aa5
aa1
aa2
aa4
aa3

5-tRNA

U G A
3-tRNA 4-tRNA

G A A G C U
G C U A C U U C G A A C U
mRNA
 aa1 peptide bonds aa5
aa2
aa3
aa4

5-tRNA

3-tRNA U G A
G A A 4-tRNA

G C U
G C U A C U U C G A A C U
mRNA

Ribosomes move over one codon

 aa5
aa4 aa199 Termination
aa3 primary aa200
structure
aa2 of a protein

aa1
terminator
200-tRNA
or stop
codon
A C U C A U G U U U A G
mRNA
 End Product
• The end products of protein synthesis is a
primary structure of a protein.
protein
• A sequence of amino acid bonded together
by peptide bonds.
bonds
aa5
aa3 aa4
aa2 aa199

aa1 aa200
 Polyribosome
• Groups of ribosomes reading same mRNA
simultaneously producing many proteins
(polypeptides).

incoming
large
subunit

1 2 3 4 5 6 7
mRNA

incoming
small subunit polypeptide
 Question:
• The anticodon UAC belongs to a tRNA that
recognizes and binds to a particular amino
acid.

• What would be the DNA base code for this

amino acid?
 Answer:
• tRNA - UAC (anticodon)
• mRNA - AUG (codon)
• DNA - TAC
DNA Replication
 Question:
• When and where does DNA Replication
take place?
 Synthesis Phase (S phase)
• S phase in interphase of the cell cycle.
• Nucleus of eukaryotes
S
DNA replication takes phase
place in the S phase.
G1 interphase G2

Mitosis
-prophase
-metaphase
-anaphase
-telophase
DNA REPLICATION CHROMOSOME
after DNA
replication
CHROMATIDS

CENT
ROME
RE
 Question:
• What is a chromosome?
 Answer:
• A chromosome is made up of a DNA -
histone protein complex called chromatin.
chromatin
• Chromatin is a long, thin fiber that is folded
and coiled to form chromosomes.
chromosomes
DNA double helix

Histone
proteins
chromosome
 Question:
• What is a replicated chromosome?
 Answer:
• A replicated chromosome consist of two
strands of identical chromosomal material
called chromatids (sister chromatids).
chromatids

chromosome

S phase: chromosomes replicate

chromatid
chromosome
chromatid
centromere
 Question:
• When is a chromatid a chromatid?
 Answer:
• A chromatid is a chromatid as long as it is
held in association with a sister chromatid at
the centromere.
centromere

centromere
chromatid
chromosome

chromatid
 DNA Replication

OLD
+
NEW
 DNA Replication
• Origins of replication

1. Replication Forks:
Forks hundreds of Y-shaped
regions of replicating DNA molecules
3’
where new strands are growing.

5’ Parental DNA Molecule Replication

Fork
3’

5’
Some of enzymes involved in DNA Replication

HELICASE
SINGLE STRAND DNA
BINDING PROTEINS
TOPOISOMERASE
DNA PRIMASE

DNA POLYMERASE
DNA LIGASE
 DNA Replication
• Strand Separation:
Separation

1. Helicase:
Helicase enzyme which catalyze the
unwinding and separation (breaking H-Bonds)
of the parental double helix.

2. Single-Strand Binding Proteins:

Proteins proteins
which attach and help keep the separated
strands apart.
 DNA Replication
• Strand Separation:
Separation
3. Topoisomerase:
Topoisomerase enzyme which relieves
stress on the DNA molecule by allowing
free rotation around a single strand.
Enzyme Enzyme

DNA
 DNA Replication

• Priming:
1. RNA primers:
primers before new DNA strands can
form, there must be small pre-existing
primers (RNA) present to start the addition of
new nucleotides (DNA Polymerase).
Polymerase)

2. Primase:
Primase enzyme that polymerizes
(synthesizes) the RNA Primer.
 DNA Replication
• Synthesis of the new DNA Strands:

1. DNA Polymerase:
Polymerase with a RNA primer in
place, DNA Polymerase (enzyme) catalyzes
the synthesis of a new DNA strand in the 5’
to 3’ direction.
direction

5’ 3’

RNA
5’
DNA Polymerase Primer
Nucleotide
 5’ reads 3' to 5'
and
STR ING

3’
AND

synthesizes in
LEA AND
STR
G

5' to 3' direction

LAG

DIN
3’
G

http://www.ncc.gmu.edu/dna/repanim.htm

http://www.bioteach.ubc.ca/TeachingResources/MolecularBiology

5’
 DNA Replication
• Synthesis of the new DNA Strands:
2. Leading Strand:
Strand synthesized as a
single polymer in the 5’ to 3’ direction.
direction

5’ 3’
5’
RNA
Nucleotides DNA Polymerase Primer
 DNA Replication
• Synthesis of the new DNA Strands:
3. Lagging Strand:
Strand also synthesized in
the 5’ to 3’ direction,
direction but discontinuously
against overall direction of replication.
Leading Strand
5 3’
’
3’ 5’
DNA Polymerase RNA Primer
5’ 3’

3’ 5’
Lagging Strand
 DNA Replication
• Synthesis of the new DNA Strands:
4. Okazaki Fragments:
Fragments series of short
segments on the lagging strand.

DNA
Okazaki Fragment Polymerase
RNA
Primer
5’ 3’

3’ 5’
Lagging Strand
 DNA Replication
• Synthesis of the new DNA Strands:
5. DNA ligase:
ligase a linking enzyme that
catalyzes the formation of a covalent
(phosphodiester) bond between the
3’ hydroxyl group at the end of one Okazaki
fragment to 5’ phosphate group of the adjacent
Okazaki fragmant.
Example: joining two Okazaki fragments together.
DNA ligase
Okazaki Fragment 1 Okazaki Fragment 2
5’ 3’

3’ Lagging Strand
5’
 DNA Replication
• Synthesis of the new DNA Strands:

6. Proofreading:
Proofreading initial base-pairing errors are
usually corrected by DNA polymerase.
polymerase
 DNA Repair
• Excision repair:

1. Damaged segment is excised by a repair

enzyme (there are over 50 repair enzymes).

2. DNA polymerase and DNA ligase replace

and bond the new nucleotides together.
DNA polymerase mistakes
= MUTATIONS =

SICKLE CELL
ANEMIA

GLUTAMATE

VALINE
 Question:
• What would be the complementary
DNA strand for the following DNA
sequence?

DNA 3’- GCGTATG - 5’

 Answer:
OLD DNA 3’- GCGTATG - 5’
NEW DNA 5’- CGCATAC - 3’
DNA Fingerprinting (Polymorphisms)
SNP = Single Nucleotide Polymorphism
 DNA Fingerprinting
Polymorphisms (SNP)

cleavage of DNA by
suspect’s blood extracted DNA restriction enzymes

A B sample

separation of DNA fragments

By electrophoresis
&
Compare with suspect’s DNA
POLYMERASE CHAIN REACTION
PCR
Kary Mullis (1993)

DNA polymerase
DNA polymerase
 GENETIC ENGINEERING
1) GENE SPLICING

Transformation

genetically engineered
“INSULIN”
 GENETIC ENGINEERING
2) GENE THERAPY

http://history.nih.gov/exhibits/genetics/sect4.htm
 ADA Cystic Fibrosis
Adenosine Deaminase
Immune system deficiency

patient lymphocytes
introduced the gene for ADA

patient epithelial cells

introduced the gene for CFTR
 3) TRANSGENIC ANIMAL OR PLANT
ALBA THE GREEN FLUORESCENT BUNNY

In one technique, embryonic stem cells (ES cells) are transformed with the desired gene (DNA).
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/T/TransgenicAnimals.html
France, 2000 Eduardo Kac –The use of genetic engineering to transfer natural or synthetic genes
to an organism, to create unique living beings. "Alba", the green fluorescent bunny, is an albino
rabbit. This means that, since she has no skin pigment, under ordinary environmental conditions
she is completely white with pink eyes. Alba is not green all the time. She only glows when
illuminated with the correct light. When (and only when) illuminated with blue light (maximum
excitation at 488 nm), she glows with a bright green light (maximum emission at 509 nm).
http://www.ekac.org/gfpbunny.html#gfpbunnyanchor