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Lecture 4

Structure and function of

nucleic acid

Contents

1. Composition of nucleic acid

2. Structure and function of
DNA
3. Structures and functions of
RNA
4. Properties of nucleic acid

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Brief history
• 1869: isolated DNA from salmon sperm (Friedrich
Miescher)
• 1944: proved DNA is genetic materials (Avery et al.)
• 1953: discovered DNA double helix (Watson and
Crick)
• 1968: decoded the genetic codes (Nirenberg)
• 1981: invented DNA sequencing method (Gilbert and
Sanger)
• 1987: launched the human genome project
• 2001: accomplished the draft map of human genome

Deoxyribonucleic acid, DNA

Nucleic acid
Ribonucleic acid, RNA

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1. The components of DNA and RNA

•DNA and RNA are polymers of nucleotide

units.
• DNA (RNA) consists of 4 kinds of
ribonucleotide units linked together through
covalent bonds.
• Each nucleotide unit is composed of
a nitrogenous base
a pentose sugar
a phosphate group

1.1 Bases
• Purines :
– Adenine (A)
– Guanine (G)
• Pyrimidines :
– Cytosine (C)
– Uracil (U)
– Thymine (T)

DNA: A,G,C,T
RNA: A,G,C,U
Thymine (T) is a 5-methyluracil (U)

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1.2 Ribose (in RNA) and deoxyribose (in DNA)

•

• Ribose and deoxyribose predominantly

exist in the cyclic form.

1.3 Nucleosides =ribose/deoxyribose + bases

•The bases are covalently attached to the 1’ position
of a pentose sugar ring, to form a nucleoside

Glycosidic bond
1

R Ribose or 2’-deoxyribose

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Adenosine, guanosine, cytidine, thymidine, uridine

1.4 Nucleotides = nucleoside + phosphate

•A nucleotide is a nucleoside with one or more phosphate
groups bound covalently to the 3’-, 5’, or ( in
ribonucleotides only) the 2’-position. In the case of 5’-
position, up to three phosphates may be attached.
Phosphate ester bonds

Deoxynucleotides Ribonucleotides
(containing deoxyribose) (containing ribose)

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BASES NUCLEOSIDES NUCLEOTIDES

Adenine (A) Adenosine Adenosine 5’-triphosphate (ATP)
Deoxyadenosine Deoxyadenosine 5’-triphosphate
(dATP)
Guanine (G) Guanosine Guanosine 5’-triphosphate (GTP)
Deoxyguanosine Deoxy-guanosine 5’-triphosphate
(dGTP)
Cytosine (C) Cytidine Cytidine 5’-triphosphate (CTP)
Deoxycytidine Deoxy-cytidine 5’-triphosphate
(dCTP)
Uracil (U) Uridine Uridine 5’-triphosphate (UTP)

Thymine (T) Thymidine/ Thymidine/deoxythymidie

Deoxythymidie 5’-triphosphate (dTTP)

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phosphate
nucleic acid nucleotides pentose
nucleosides
bases

NH2

N
O
HO P O CH2 N O
O
OH

OH OH

Composition of DNA and RNA

Nucleic
base ribose
acid

DNA A、G、C、T deoxyribose

RNA A、G、C、U ribose

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1.5 Some important nucleotides

• dATP, dGTP, dCTP, dUTP
– Raw materials for DNA biosynthesis.
• ATP, GTP, CTP, UTP
– Raw materials for RNA biosynthesis
– Energy donor
– Important co-enzymes
• Cycling nucleotides—cAMP, cGMP
– Secondary messengers in hormones action.

Nucleic acid derivatives

Multiple phosphate nucleotides
adenosine monophosphate (AMP)
adenosine diphosphate (ADP)
adenosine triphosphate (ATP)

NNHH22
NH2
N
N
N NN
O OO N
O O O
N
H O P HOO PP OO C
CH 2 N NN
HO P O P O P O CH2 O N N
OH OOHH O
OH OH OH

ADP AMP OOHH OH

H
OH
ATP

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2. Structure and function of DNA

2.1 Primary structure

 Definition: the base sequence (or the
nucleotide sequence) in
polydeoxynucleotide chain.
 The smallest DNA in nature is virus DNA.
The length of φX174 virus DNA is 5,386
bases (a single chain).
 The DNA length of human genome is
3,000,000,000 pair bases.

5’end

Phosphodiester
bond

3’ end: free hydroxyl

(-OH) group

• 3’,5’ phosphodiester bond link nucleotides

together to form polynucleotide chains

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The structure of a DNA chain can be

concisely represented

• An even more abbreviated notation for

this chain is
– pApCpGpTpA
– pACGTA
• The base chain is written in the 5’ →3’
direction

2.2 Secondary structure

The secondary structure is defined as

the relative spatial position of all the
atoms of nucleotide residues.

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Secondary structure
— DNA double helix structure

Francis H.C. Crick

•Watson and Crick , 1953
•The genetic material of
all organisms except for
some viruses.
•The foundation of the
molecular biology.

James D. Watson

The discovery of DNA double helix

• Chargaff's Rule
(A=T, G=C in DNA)

• Franklin, Wilkins:
X-ray
Diffraction
Refined Structure

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Molarity of bases

A G C T A/T G/C G+C Pu/Py

E. coli 26.0 24.9 25.2 23.9 1.09 0.99 50.1 1.04

Tuberc 15.1 34.9 35.4 14.6 1.03 0.99 70.3 1.00

ulosis
Yeast 31.7 18.3 17.4 32.6 0.97 1.05 35.7 1.00

Cow 29.0 21.2 21.2 28.7 1.01 1.00 42.4 1.01

Pig 29.8 20.7 20.7 29.1 1.02 1.00 41.4 1.01

Human 30.4 19.9 19.9 30.1 1.01 1.00 39.8 1.01

DNA double helix

Essential for replicating DNA
•Two separate strands and transcribing RNA
•Antiparellel (5’3’
direction) 3’
5’
•Base pairing:
hydrogen bonding
that holds two
strands together
•Complementary
(sequence)

• Sugar-phosphate
backbones (negatively
charged): outside
• Base pairs (stack one
above the other): inside 3’
5’

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4
32 1
6
87 5 1
9 4
3 2

A:T G:C

Base pairing

B form of DNA double

helix
• Right-handed helix;
•The diameter of the
double helix：2 nm
• The distance
between two base
pairs: 0.34 nm;
• Each turn of the
helix involves 10 bases
pairs, 3.4 nm.

 Stable configuration
can be maintained by
hydrogen bond and base
stacking force
(hydrophobic interaction).

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Groove binding

• Small molecules like drugs bind in the minor

groove, whereas particular protein motifs can
interact with the major grooves.

• Watson, Crick, and Wilkins

shared the Nobel Prize in
medicine or physiology in 1962
for this brilliant accomplishment.

• The discovery of the DNA

double helix revolutionized
biology: it led the way to an
understanding of gene function
in molecular terms (their work is
recognized to mark the
beginning of molecular biology).

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Conformational variation in
double-helical structure
• B-DNA
• A-DNA
• Z-DNA

• B-form: the duplex structure proposed by Watson and Crick is

referred as the B-form DNA.
•It is the standard structure for DNA molecules.
•A-form: at low humidity the DNA molecule will take the A-form:
•The A-form helix is wider and shorter, with a shorter more compact
helical structure, than the B-form helix.
• Z-form: the Z-form DNA is adopted by short oligonucleotides.
•It is a left-handed double helix in which backbone phosphates zigzag.

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2.3 Tertiary structure :

• Supercoils: double-stranded circular DNA
form supercoils if the strands are
underwound (negatively supercoiled) or
overwound (positively supercoiled).

Increasing degree of supercoiling

Relaxed supercoiled

• If the strands
are overwound,
form positively
supercoiled;
• If the strands
are underwound,
form negatively
supercoiled.

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• The DNA in a prokaryotic cell is

a supercoil.

• Supercoiling makes the DNA molecule more

compact thus important for its packaging
in cells.

2.4 Eukaryotic DNA

• DNA in eukaryotic cells is highly

packed.
• DNA appears in a highly ordered form
called chromosomes during metaphase,
whereas shows a relatively loose form
of chromatin in other phases.
• The basic unit of chromatin is
nucleosome.
• Nucleosomes are composed of DNA
and histone proteins.

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Nucleosome
• The chromosomal DNA
is complexed with five
types of histone.
•H1, H2A, H2B, H3 and
H4.
•Histons are very basic
proteins, rich in Arginine
and Lysine.

•Nucleosomes: regular association of DNA with

histones to form a structure effectively compacting
DNA. ”beads”

Beads on a string
• 146 bp of
negatively
supercoiled DNA
winds 1 ¾ turns
around a histone
octomer.
• H1 histone binds
to the DNA
spacer.

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The importance of packing of DNA into

chromosomes

 Chromosome is a compact form of the DNA

that readily fits inside the cell
 To protect DNA from damage
 DNA in a chromosome can be transmitted
efficiently to both daughter cells during
cell division
 Chromosome confers an overall organization
to each molecule of DNA, which facilitates
gene expression as well as recombination.

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2.5 Functions of DNA

• The carrier of genetic information.
• The template strand involved in replication
and transcription.

Gene: the minimum functional unit in DNA

Genome: the total genes in a living cell or

living beings.

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3. Structures and functions of RNA

Conformational variability of RNA is important

for the much more diverse roles of RNA in
the cell, when compared to DNA.
Types :
• mRNA: messenger RNA, the carrier of genetic
information from DNA to translate into protein
• tRNA: transfer RNA , to transport amino acid to
ribosomes to synthesize protein
• rRNA: ribosomal RNA, the components of
ribosomes
• hnRNA: Heterogeneous nuclear RNA
• snRNA: small nuclear RNA

Classes of eukaryotic RNAs

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RNA structure
• RNA molecules are largely single-
stranded but there are double-
stranded regions.

3.1 Messenger RNA( mRNA)

• Function: the carrier of genetic
information from DNA for the
synthesis of protein.
• Comprises only about 5% of the
RNA in the cell.
• Composition: vary considerably in
size (500-6000 bases in E. coli)

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Eukaryotic mRNA Structure

(1) Capping: linkage of 7-
methylguanosine to the 5’ terminal
residue.
(2) Tailing: attachment of an
adennylate polymer (poly A, 20~250
nucleotides) at the 3’ terminal.

3.2 Transfer RNA (tRNA)

• They make up 15% of the RNA in the cell.
• Function: Transport amino acids to ribosomes for
assembly into proteins.
• There are at least 20 types of tRNA in one cell.

• Primary Structure :
– 74~95 bases, the smallest of the three major
RNA.
– Modified bases: pseudouridine (ψ)
methylguanosine
dihydrouridine (D)
– The sequence CCA at the 3’ terminus

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Secondary structure: cloverleaf

• Four loops and four
arms
– Amino acid arm
(7bp): to bide amino
acid
– D loop(8-14bp) and
D arm(3-4bp):
– Anticoden loop(5bp)
and arm(7bp): to
recognize amino acid
coden on the mRNA.
– TψC loop（7bp） and
arm(5bp)
– Variable loop(4-5bp
or 13-21bp)

•Tertiary structure of tRNA

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3.3 Ribosomal RNA (rRNA)

• Components of ribosomes.
• They make up 80% of the RNA in the cell.
* The species of rRNA
•Eukaryotes •Prokaryotes
• 5S rRNA • 5S rRNA
• 28S rRNA • 23S rRNA
• 18S rRNA • 16S rRNA
• 5.8S rRNA
• S represents Svedberg units, they represent
measures of sedimentation rate.

The proposed
secondary structure
for E.coli 16S rRNA

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Ribosomes
• Ribosomes are cytoplasmic structures that
synthesize protein, composed of RNA (2/3)
and protein (1/3).
• The ribosomes of prokaryotes and
eukaryotes are similar in shape and function.
The difference between them is the size
and chemical composition.

Three rRNA
52 proteins

Four rRNA
83 proteins

• Ribosomes are ribonucleoprotein particles for

synthesizing proteins.

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• The structure of the

complete 70S ribosome
containing mRNA and
two or three tRNAs was
solved by x-ray
crystallography to a
resolution of 5.5 Å.
(Cate et al., 1999; Yusupov et
al., 2001).

Other RNAs
• Small nuclear RNA (snRNA)
– Involved in mRNA processing
• Small nucleolar RNA (snoRNA)
– Play a key role in the processing of rRNA
molecules
• Small cytoplasmic RNA (scRNA)
– Involved in the selection of proteins for export
• Catalytic RNA or Ribozyme
• Small interfering RNA (siRNA)
– Interfere with the expression of a specific
gene
• RNomics

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4. Physical and Chemical

Properties of Nucleic Acids

General properties
• Acidity
– Amphiphilic molecules; normally acidic because
of phosphate.
• Viscosity
– Solid DNA: white fiber; RNA: white powder.
Insoluble in organic solvents, can be precipitate
by ethanol.
• Optical absorption
– UV absorption due to aromatic groups.
• Thermal stability
– Disassociation of dsDNA (double-stranded DNA)
into two ssDNAs (single-stranded DNA).

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4.1 UV Absorption
• Specific absorption at 260nm.
• This can be used to identify nucleic
acid.

The UV absorption spectra of the common ribonucleotides

4.2 Denaturation

• Concept:
• The course of hydrogen bonds broken,
3-D structure was destroyed, the double
helix changed into single strand irregular
coil.
• Results:
(1) the value of 260nm absorption is increased;
(2) biological functions are lost.

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• Heat denaturation and Tm

• When DNA were

heated to certain
temperature, the
absorption value at
260nm would increased
sharply，which indicates
that the double strand
helix DNA was
separated into single
strand.

•Tm (melting temperature of DNA):

• The temperature of UV absorption increase to an
half of maximum value in DNA denaturation.

• Factors affect Tm:

G-C content:
•There are three hydrogen bonds between G-C
pair. The more G-C content, the higher Tm value.

Less G+C Tm of
Higher G+C two DNA
molecules with
different G+C
content

Temperature

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4.3 Renaturation of DNA

• When slowly cooling down (Annealing)
the denatured DNA solution, the single
strand DNA can reform a double strands
helix to recover its biological functions.

Molecule hybridization
• During the course of
lowing down denaturing
temperature, between
different resource DNAs
or single stand DNA and
RNA with
complementary bases
will repair into a double
strands to form a hybrid
DNA or DNA-RNA . This
course is called molecule
hybridization.

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Points
• The components of DNA and RNA
– Nucleotide: base (A,G,C,T,U), pentose sugar
(Ribose and deoxyribose), phosphate group
• Structure and function of DNA
– Primary structure: 3’,5’ phosphodiester bond
– Secondary structure: DNA double helix
– Tertiary structure: supercoil
– Eukaryotic chromosomes: nucleosome
• Structures and functions of RNA
– mRNA, tRNA, rRNA
• Properties of nucleic acid
– UV absorption, denaturation and renaturation,
molecule hybridization

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