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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.

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Presentation on theme: "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."— Presentation transcript:

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 N.B. Some images used in these slides are from the textbooks listed and are not covered under the Creative Commons license as yet 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. To view a copy of this license, visit or send a letter to Creative Commons, 171 Second Street, Suite 300, San Francisco, California 94105, USA.http://creativecommons.org/licenses/by-nc-sa/2.0/uk/

2 learning objectives 1)Compare sizes of DNA and understand the C-value paradox 2)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 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 Outline

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

5 genome sizes organism Number of base pairs (kb) viruses Lambda bacteriophage ( λ)48.6 bacteria Eschericia coli4,640 eukaryotes Yeast13,500 Drosophila165,000 Human3.3 x 10 6

6 What is the Human genome project? 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 Goal: to sequence the entire human nuclear genome

7 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? nuclear DNA Hair root mitochondrial DNA Hair shaft....or in forensics

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

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

15 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 Explaining the paradox

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 Supercoiled DNA moves faster than relaxed DNA Allows packing / unpacking of DNA

19 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 Supercoiling topology No supercoiling (left) to tightly supercoiled (right)

20 Visualising DNA/RNA with dyes Ethidium bromide EBr

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

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

23 Forces stabilising nucleic acid structures Applications in polymerase chain reaction (PCR) A) Denaturation and renaturation of DNA 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

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 ~ 25 o C below Tm Property used in PCR and hybridisation techniques

28

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

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

31 Forces stabilising nucleic acid structures C) Base pairing

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

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 1)mRNA: directs the ribosomal synthesis of polypeptides and other types of RNA (translation) 2)Ribosomal RNA: have structural & functional roles 3)Transfer RNA: deliver amino acids during protein synthesis 4)Ribonucleoproteins: take part in post transcriptional processing

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

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

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