Chapter 8

Nucleotides and Nucleic Acids

 Learning Objectives
1. Summarize the central dogma of molecular biology, and cite exceptions to the
original model.
2. Compare and contrast prokaryotic and eukaryotic gene structure.
3. Compare and contrast the structure of DNA and RNA, explaining the difference
between the constituent bases, sugars, nucleosides and nucleotides.
4. Identify and draw the structures of DNA and RNA bases plus nucleosides,
nucleotides and polynucleotides including ribose and deoxyribose
5. Compare and contrast the different types of RNA.
6. Integrate the terminology and defining structural features that distinguish different
classes of nucleotide metabolites (such as purine vs. pyrimidine, bases vs.
nucleoside vs. nucleotide, and ribo- vs. deoxyribo-).
7. Describe the double-stranded, helical, and antiparallel chain structure of DNA and
how it relates to the processes of DNA replication, transcription, recombination and
repair.
8. Describe how DNA and DNA processes can be used as therapeutic targets (e.g.
anticancer and antibacterial drugs).
9. Use the genetic code to predict the amino acid sequence of a protein for a given
nucleic acid sequence and demonstrate how nucleotide mutations can lead to
alterations in the primary structure of a protein.
Exam Resources: Lehninger’s Chapter 8, Class Notes.
 Chapter Overview

I. Nucleotides and Nucleic Acids

II. DNA Structure
III. DNA Analysis
IV. RNA Structure
 Functions of
I. Nucleotides and Nucleic Acids
• Nucleotide Functions:
– Energy for metabolism (ATP)
– Enzyme cofactors (NAD+)
– Signal transduction (cAMP)

• Nucleic Acid Functions:

– Storage of genetic info (DNA)
– Transmission of genetic info (mRNA)
– Processing of genetic information (ribozymes)
– Protein synthesis (tRNA and rRNA)
 Nucleotides and Nucleosides
• Purines (A/G), Pyrimidines (C/T/U)
• Nucleotide =
– Nitrogeneous base
– Pentose
– Phosphate
• Nucleoside =
– Nitrogeneous base
– Pentose
• Nucleic acids built from dNTP
 Adenosine triphosphate (ATP)

Adenine

γ β α
Phosphates
Ribose

Guanosine triphosphate (GTP)

Guanine

γ β α
Phosphates

Ribose
 Purine Bases
• Adenine and guanine are found in both RNA and DNA
• Also good H-bond donors and acceptors
• Neutral molecules at pH 7
 Pyrimidine Bases
• Cytosine is found in both DNA and RNA
• Thymine is found only in DNA
• Uracil is found only in RNA
• All are good H-bond donors and acceptors
• Neutral molecules at pH 7
 DNA bases built from amino acid
precursors

CTP/TTP

UTP

ATP/GTP
Classify each structure

Base

Base
 Tautomerism of Nucleobases
• Prototropic tautomers are structural isomers that differ in the location of protons
• Keto-enol tautomerism is common in ketones
• Lactam-lactim tautomerism occurs in some heterocycles
• Both tautomers exist in solution but the lactam forms are predominant at neutral pH

Adenine, Cytosine Guanine, Thymine

Nucleotides
 Ribose
• -D-ribofuranose in RNA, -2’-deoxy-D-
ribofuranose in DNA
• Different puckered conformations of
the sugar ring are possible
• Rotation around six bonds
 Conformation around N-Glycosidic Bond

• In nucleotides the pentose ring is attached to the nucleobase via N-glycosidic

bond
• Relatively free rotation can occur around the N-glycosidic bond in free nucleotides
• Angle near 0 corresponds to syn conformation
• Angle near 180 corresponds to anti conformation
• Anti conformation is found in normal B-DNA
 Minor Nucleosides in DNA
• Modification is done after DNA
synthesis
• 5-Methylcytosine is common in
eukaryotes, also found in bacteria
• N6-Methyladenosine is common in
bacteria, not found in eukaryotes
• Epigenetic marker
• Way to mark own DNA so that
cells can degrade foreign DNA
(prokaryotes)
• Way to mark which genes should
be active (eukaryotes)
 Minor Nucleosides in RNA

• Inosine sometimes found in the “wobble

position” of the anticodon in tRNA
– Made by de-aminating adenosine
– Provides richer genetic code

• Pseudouridine ( ) found widely in tRNA

and rRNA
– More common in eukaryotes but
found also in eubacteria
– Made from uridine by enzymatic
isomerization after RNA synthesis
– May stabilize the structure of tRNA
– May help in folding of rRNA
Other Functions of Nucleotides:
Energy, Coenzymes, Regulatory

(vitamin B5)

(vitamin B2)

(vitamin B3)
 Polynucleotides
• Covalent bonds formed via
phosphodiester linkages
– negatively charged backbone
• DNA backbone is fairly stable
– Ancient DNA
– Hydrolysis accelerated by enzymes
(DNAse)
• RNA backbone is unstable
– In water, RNA lasts for a few years
– In cells, mRNA is degraded in few
hours
• Linear polymers
– No branching or cross-links
• Directionality
– 5’ end is different from 3’ end
– We read the sequence from 5’ to 3’ note: Nucleotides are joined by
3’- 5’-phosphodiester bonds
 Hydrolysis of RNA
• RNA is unstable under alkaline
conditions
• Hydrolysis is also catalyzed by
enzymes (RNase)
• RNase enzymes are abundant:
– S-RNase in plants prevents
inbreeding
– RNase P is a ribozyme (enzyme
made of RNA) that processes
tRNA precursors
– Dicer is an enzyme that cleaves
double-stranded RNA into
oligonucleotides
• protection from viral
genomes
• RNA interference technology
– Ribosome
 Hydrogen-Bonding Interactions
• Two bases can hydrogen bond to form a base pair
• For monomers, large number of base pairs is possible
• Watson-Crick base pairs predominate in double-stranded DNA (A/T, G/C)
• Purine pairs with pyrimidine

A-T base pairs have 2 hydrogen bonds

G-C base pairs have 3 hydrogen bonds

II.
• Double helix
DNA Structure

• Sugar/phosphate backbone (– charge)

• Major/minor groove
• Deoxyribose
• Base pairing
• Antiparallel
• 34 A / 10.5 bp/turn
• 2 nm wide
 10.5 bp/turn
DNA Structure
in DNA
- Major groove is wide, easily accessible to proteins
- Minor groove is narrow
 DNA Stability:
H-Bonding & Base Stacking
 Replication of Genetic Code
• Strand separation occurs first
• Each strand serves as a template
for the synthesis of a new strand
• Synthesis is catalyzed by
enzymes known as DNA
polymerases
• Newly made DNA molecule has
one daughter strand and one
parent strand
 Molecular Mechanisms of
Spontaneous Mutagenesis
• Half a million/day/cell
• Deamination
– Very slow reactions
– Large number of residues
– The net effect is significant: 100 C
U events /day in a mammalian cell
• Depurination
– N-glycosidic bond is hydrolyzed
– Significant for purines: 10,000
purines lost/day in a mammalian cell
• Oxidative damage
• Alkylation
• UV, ionizing radiation
• Cells have mechanisms to correct
most of these modifications
III. DNA Analysis
• Absorption of UV light at 250–270 nm is due to   * electronic transitions
• Excited states of common nucleobases decay rapidly via radiationless transitions
– Effective photoprotection of genetic material
– No fluorescence from nucleic acids
 DNA Denaturation
• Covalent bonds remain intact
– Genetic code remains
intact
• Hydrogen bonds are broken
– Two strands separate
• Base stacking is lost
– UV absorbance increases

Denaturation can be induced by

high temperature, or change in
pH

Denaturation may be reversible:

annealing
 Thermal DNA Denaturation (Melting)
• DNA exists as double helix at normal temperatures,
strands dissociate at high temperatures
• The reversible thermal denaturation and annealing
form basis for the polymerase chain reaction
• DNA denaturation is commonly monitored by UV
spectrophotometry at 260 nm
• Tm increased by longer length, higher GC content,
higher salt

Hyperchromicity
 Two near-complementary
DNA strands can hybridize
• Detection of a specific DNA molecule in complex mixture
- radioactive detection
- fluorescent DNA chips
• Amplification of specific DNA
- polymerase chain reaction
- site-directed mutagenesis
• Evolutionary relationships
• Antisense therapy
Sanger sequencing
Automating DNA sequencing reactions
 Automated
DNA Synthesis
IV. RNA Structure

• 2’-hydroxyl
• Uracil
• Single-stranded
• Double-stranded form is
helical (A form, not B form,
due to OH), but irregular
• Not genetic material (except
for viruses)
• Adaptor between mRNA and
amino acids
• Ribosome
 RNA Structure
• Synthesized using DNA template
• Contains ribose instead of deoxyribose, uracil instead of thymine

tRNA

Hammerhead ribozyme

Self-splicing intron
 Complex shapes due to:
• H-bonds in (unusual) base pairing and stacking
• Complementary sequences pairing
• Minimum free energy conformation
 RNA Motifs as building block

E-loop motif
sarcin-ricin
kink-turn motif C-loop motif motif reverse kink-turn
motif
Figure 3. Base-pairing patterns of the query motif structures in 2D diagrams.

http://genome.ucf.edu/RNAMotifScan/ Zhong C et al. Nucl. Acids Res. 2010;38:e176-e176

Non-Watson-Crick G=U base
pairs (●) in E. coli Rnase P

G-U base pairs have

TWO hydrogen bonds
In the Ribosome:
- Protein component assists in tertiary structure & protection
- Divalent cations also assist in stabilization & shield backbone
to allow packing
 Summary of A-form RNA versus B-form DNA
• Most RNA and RNA-DNA duplexes • B-form is most common DNA
are A-form conformation in vivo
• Shorter, wider helix than B-form. • Narrower, more elongated helix than A-
• Deep, narrow major groove not form.
easily accessible to proteins • Wide major groove easily accessible to
• Wide, shallow minor groove proteins
accessible to proteins, but lower • Narrow minor groove
information content than major groove • Favored conformation at high water
• Favored conformation at low water concentrations (hydration of minor
concentrations groove seems to favor B-form)
• Base pairs tilted to helix axis and • Base pairs nearly perpendicular to helix
displaced from axis (11 bp/turn) axis (10.5 bp/turn)
• Sugar pucker C3'-endo (note: in RNA • Sugar pucker C2'-endo
2'-OH inhibits C2'-endo conformation)

Note: glycosidic bond conformation is in the

anti form for both A-form and B-form.
 Learning Objectives
1. Summarize the central dogma of molecular biology, and cite exceptions to the
original model.
2. Compare and contrast prokaryotic and eukaryotic gene structure.
3. Compare and contrast the structure of DNA and RNA, explaining the difference
between the constituent bases, sugars, nucleosides and nucleotides.
4. Identify and draw the structures of DNA and RNA bases plus nucleosides,
nucleotides and polynucleotides including ribose and deoxyribose
5. Compare and contrast the different types of RNA.
6. Integrate the terminology and defining structural features that distinguish different
classes of nucleotide metabolites (such as purine vs. pyrimidine, bases vs.
nucleoside vs. nucleotide, and ribo- vs. deoxyribo-).
7. Describe the double-stranded, helical, and antiparallel chain structure of DNA and
how it relates to the processes of DNA replication, transcription, recombination and
repair.
8. Describe how DNA and DNA processes can be used as therapeutic targets (e.g.
anticancer and antibacterial drugs).
9. Use the genetic code to predict the amino acid sequence of a protein for a given
nucleic acid sequence and demonstrate how nucleotide mutations can lead to
alterations in the primary structure of a protein.
Exam Resources: Lehninger’s Chapter 8, Class Notes.
 Chapter 8 Summary
• Function of nucleotides and nucleic acids
• Names and structures of common
nucleotides
• Structural basis of DNA function
• Reversible denaturation of nucleic acids
• Chemical basis of mutagenesis
• Automated DNA sequencing & synthesis
• RNA structure
 References
• Nelson & Cox. 2017. Lehninger Principles of Biochemistry,
7th edition by W.H. Freeman & Co.
• Lieberman & Marks. 2013, Mark’s Basic Medical
Biochemistry, 4th edition by Lippincott William & Wilkins
• Watson, Baker, Bell, Gann, Levine & Losick. 2014. Molecular
Biology of the Gene, 7th edition by Pearson.
• Baynes & Dominiczak. 2009, Medical Biochemistry, 3rd
edition by Mosby Ltd.
• White, Drummond & Fuqua. 2012. The Physiology and
Biochemistry of Prokaryotes, 4th edition by Oxford
University Press, Inc.
• Schaechter, Ingrahm & Neidhardt. 2006. Microbe by
American Society for Microbiology Press.
The following slides are useful slides that I did not present.
There are here for reference in order to help you study.
 DNA bases built from amino acid precursors

CTP/TTP

UTP
DNA bases built from amino acid precursors

ATP/GTP
 Atom numbering in Nucleosides

Deoxyribonucleoside to sugar to sugar

Ribonucleoside

to sugar to sugar to sugar

*Know structures of all

RNA/DNA bases*
 Nucleoside and nucleotide structures

Marks’ Fig 12.3

A-form RNA versus B-form DNA
 Major and Minor groove
Major Groove Minor Groove
Adenine N6 amino, N7 Adenine N3
Cytosine N4 amino Cytosine C2 carbonyl Guanine
Guanine C6 carbonyl, N7 N2 amino, N3
Thymine C4 carbonyl, C5 methyl Thymine C2 carbonyl
Uracil C4 carbonyl, N7 Uracil C2 carbonyl

to sugar to sugar
to sugar to sugar
 A-form RNA versus B-form DNA
• Most RNA and RNA-DNA duplexes • B-form is most common DNA
are A-form conformation in vivo
• Shorter, wider helix than B-form. • Narrower, more elongated helix than A-
• Deep, narrow major groove not form.
easily accessible to proteins • Wide major groove easily accessible to
• Wide, shallow minor groove proteins
accessible to proteins, but lower • Narrow minor groove
information content than major groove • Favored conformation at high water
• Favored conformation at low water concentrations (hydration of minor
concentrations groove seems to favor B-form)
• Base pairs tilted to helix axis and • Base pairs nearly perpendicular to helix
displaced from axis (11 bp/turn) axis (10.5 bp/turn)
• Sugar pucker C3'-endo (note: in RNA • Sugar pucker C2'-endo
2'-OH inhibits C2'-endo conformation)

Note: glycosidic bond conformation is in the

anti form for both A-form and B-form.
Non-Watson-Crick base pair:
G-U base pair in RNA

G-U base pairs have 2 hydrogen bonds

Molecular Mechanisms of Oxidative
and Chemical Mutagenesis
• Oxidative damage
• hydroxylation of guanine
• mitochondrial DNA is most susceptible

• Chemical alkylation
• methylation of guanine

• Cells have mechanisms to correct most of

these modifications.
 Clinical Correlation: Viruses
• DNA viruses:
– Viruses possess double-stranded DNA
• e.g. adenovirus, small pox virus, herpes simplex virus, human papilloma viruses &
Epstein-Barr virus
– Viruses possess single-stranded DNA
• Such as Torque teno virus & Parvovirus B19
• RNA viruses:
– Viruses possess double-stranded RNA genomes
• e.g. rotavirus & reovirus. These genomes are always segmented.
– Viruses possess positive-sense single-stranded RNA genomes.
• positive-sense: similar to host mRNA, so can be immediately translated
• e.g. picornaviruses (Hepatitis A virus, enteroviruses, rhinoviruses, Norovirus,
poliovirus, and foot-and-mouth virus), SARS virus, Zika virus hepatitis C virus, yellow
fever virus & rubella virus plus the Chikungunya virus.
– Viruses possess negative-sense single-stranded RNA genomes.
• negative-sense: 3‘-5‘ viral RNA, so must be converted to 5’-3’ mRNA first
• The deadly Ebola and Marburg viruses as well as influenza virus, measles, mumps
and rabies.
 Nucleotide
Analogs as Drugs
*
for DNA viruses

Approved antivirals for DNA *

viruses - Hepatitis B virus and
Herpes virus infections.
(a)–(b) are approved for
HBV (hepatitis B virus)
(c)–(f) are approved for HSV
(herpes simplex virus) infections
(g)–(j) are approved for HCMV
(human cytomegalovirus)
infections
(k) for VZV (Varicella-zoster
virus) infections.