1. Properties and functions of nucleic acids
These slides provides an overview of some of the properties of nucleic acids (DNA and RNA) and its applications in molecular biology
Dr. Momna Hejmadi, University of Bath
DNA basics resources created by Dr. Momna Hejmadi, University of Bath, 2010, is licensed under the Creative Commons Attribution-Non-Commercial-Share Alike 2.0 UK: England & Wales License.
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N.B. Some images used in these slides are from the textbooks listed and are not covered under the Creative Commons license as yet
N.B. Some images used in these slides are from the textbooks listed and are not covered under the Creative Commons license as yet

2. learning objectives Reference
Compare sizes of DNA and understand the C-value paradox
Understand the Human Genome Project
3) Be able to describe how the different helical topologies of DNA contribute to packing?
4) Understand the factors that contribute to the stability of the DNA double helix?
5) Appreciate the diverse functions of nucleic acids
Reference
Chapter 29: Biochemistry (3e) by D Voet and J Voet
(Wiley Publishing)

3. Outline Genomes and the Human Genome project C-value paradox
DNA topology and function
Factors that stabilise DNA
a) denaturation and renaturation
b) Sugar-phosphate chain conformations
c) Base pairing and base stacking
d) hydrophobic and ionic interactions
Functions of nucleic acids

4. DNA vs RNA size
DNA molecules tend to be larger than RNA molecules

5. genome sizes organism Number of base pairs (kb) viruses
organism Number of base pairs (kb)
viruses
Lambda bacteriophage ( λ) 48.6
bacteria
Eschericia coli 4,640
eukaryotes
Yeast ,500
Drosophila ,000
Human x 106

6. What is the Human genome project?
Goal: to sequence the entire human nuclear genome
Public consortium
Headed by F Collins
Started in mid 80’s
Working draft completed in 2001
Nature: Feb 2001
Celera Genomics
Headed by C Venter
Started in mid 90’s
Working draft completed in 2001
Science: Feb 2001
Human genome = 3.3 X 10 9 base pairs
Number of genes = 26 – 32,000 genes

7. Mitochondrial genome (16.6kbp) – multicopy, circular, ds DNA
The human genome
Nuclear genome (3.2 Gbp)
24 types of linear chromosomes
Y- 51Mb and chr1 -279Mbp
~ 30,000 genes
Mitochondrial genome (16.6kbp) – multicopy, circular, ds DNA
Gene and gene-related
Everything else

8. Why do we need the DNA blueprint?
Individual human variation is 0.1%
i.e. 1.4 million sequence variations
Applications in medicine, forensics, bioarchaeology, anthropology, human evolution, human migration etc

9. ....in disease

10. ……or risk of disease
N(291)S

11. ....or in pharmacogenomics

12. what can a single human hair tell you?
....or in forensics
what can a single human hair tell you?
mitochondrial DNA
Hair shaft
nuclear DNA
Hair root

13. Does size matter? C-value paradox
Boa constrictor
Genome size: 2.1 Gbp
Homo sapiens sapiens
Genome size: 3.2 Gbp
mountain grasshopper
Podisma pedestris
Genome size: 18 Gbp
protozoan
Amoeba dubia
Genome size: 670Gbp
C value: DNA content of any haploid cell

14. Comparative genome sizes
C-value paradox
Why is there a discrepancy between
genome size and
genetic complexity?

15. Protein domains contribute to organism complexity
Explaining the paradox
Genome sizes vary due to the presence of repetitive DNA
Repetitive DNA families constitute nearly one-half of genome (~45%)
Protein domains contribute to organism complexity

16. Largest known mammalian gene is….
DMD gene 2.5 Mbp (0.1% of the genome)
Mutations cause Duchenne’s muscular dystrophy
characterized by rapid progression of muscle degeneration which occurs early in life.
‘scoliosis’

17. Duchenne’s muscular dystrophy
Mutations in DMD gene lead to non functional dystrophin protein (localised on periphery of normal muscle fibres)
DMD patient
Normal

18. Topology of DNA DNA supercoiling: coiling of a coil
Important feature in all chromosomes
Allows packing / unpacking of DNA
Supercoiled DNA moves faster than relaxed DNA

19. Supercoiling topology
No supercoiling (left) to tightly supercoiled (right)
negatively supercoiled
Results from under or unwinding
Important in DNA packing/unpacking
e.g during replication/transcription
positively supercoiled
Results from overwinding
Also packs DNA but difficult to unwind

20. Visualising DNA/RNA with dyes
EBr
Ethidium bromide

21. Supercoiling explains why an uncut plasmid gives
more than one band on a gel
Full length linear
Relaxed circle
supercoiled

22. DNA supercoiling takes 2 forms
toroidal (DNA around histones) or interwound (bacterial chromosomes)

23. Forces stabilising nucleic acid structures
The forces that stabilise nucleic acids (N.As) are largely common to those that stabilise proteins
 The way they combine gives N.As very different properties
A) Denaturation and renaturation of DNA
Applications in polymerase chain reaction (PCR)

24. Denaturation of DNA Also called melting
Occurs abruptly at certain temperatures
Tm – temp at which half the helical structure is lost

25. DNA melting curve

26. Tm varies according to the GC content
High GC content - high Tm
GC rich regions tend to be gene rich

27. Renaturation of DNA Also called annealing Occurs ~ 25oC below Tm
Property used in PCR and hybridisation techniques

29. Forces stabilising nucleic acid structures
B) Sugar-phosphate chain conformations

30. The out of plane atom is usually C2’ or C3’
1. N-glycosidic linkage has only one or two stable positions (syn/anti)
Conformation determined by 7 angles ( )
2. Sugar ring puckers to relieve crowding of substituents that would otherwise occur in planar conformation
Planar
Puckered
The out of plane atom is usually C2’ or C3’
Endo conformation (same side as C5’)
B-DNA is C2’ endo
Fig: 28-18: Voet and Voet

31. Forces stabilising nucleic acid structures
C) Base pairing

32. Hoogsten base pairs stabilise tRNA tertiary structure
D. Factors that stabilise N.As (c)
(C) Base pairing
When monomeric A and T are co-crystallised:
- They form Hoogsteen geometry
Hoogsten base pairs stabilise tRNA tertiary structure
Watson-Crick geometry is preferred in double helices due to various environmental influences T A

33. Forces stabilising nucleic acid structures
D) Base stacking and hydrophobic interactions
Under aqueous conditions
Bases aggregate due to the stacking of planar molecules
This stacking is stabilised by hydrophobic forces

34. Forces stabilising nucleic acid structures
Tm of a DNA duplex increases with cationic concentration
Caused by electrostatic shielding of anionic phosphate groups
e.g. Mg 2+ more effective than Na+
E) Ionic interactions

35. Functions of nucleic acids
1) Storage of genetic information
2) Storage of chemical energy e.g. ATP
3) Form part of coenzymes
e.g. NAD+, NADP+, FAD and coenzyme A
4) Act as second messengers in signal transduction e.g. cAMP

36. Functions of nucleic acids
1) Storage of genetic information
DNA is the hereditary molecule in almost all cellular life forms. It has 2 main functions:
Replication (making 2 copies of the genome) before every cell division
Transcription: process of copying a portion of DNA gene sequence into a single stranded messenger RNA (mRNA)

37. RNA (ribonucleic acid)
Has a more varied role. 4 main types of RNA are
mRNA: directs the ribosomal synthesis of polypeptides and other types of RNA (translation)
Ribosomal RNA: have structural & functional roles
Transfer RNA: deliver amino acids during protein synthesis
Ribonucleoproteins: take part in post transcriptional processing

38. ATP (adenosine triphosphate)
Functions of nucleic acids
2) Storage of chemical energy e.g. ATP
ATP (adenosine triphosphate)
Involved in
1) Early stages of nutrient breakdown
2) Physiological processes
3) Interconversion of nucleoside triphosphates

39. Functions of nucleic acids
3) Form part of coenzymes
e.g. NAD+, NADP+, FAD and coenzyme A

40. Functions of nucleic acids
4) Act as second messengers in signal transduction e.g. cAMP (cyclic Adenosine Mono Phosphate)
Primary intracellular signalling molecule (second messenger system)
Glycogen metabolism
cAMP dependent kinase (cAPK)
Gluconeogenesis
Fatty acid metabolism - thermogenesis