MOLECULAR STRUCTURE OF DNA

Transcription

1 MOLECULAR STRUCTURE OF DNA

2 Characteristics of the Genetic Material 1. Replication Reproduced and transmitted faithfully from cell to cell (generation to generation) 2. Information Storage Biologically useful information in a stable form 3. Expression of Information Other biologically important molecules, and ultimately cells and organisms, will be produced and maintained 4. Variation (by mutation) Some change is required

3 Experiments Defining the Genetic Material Griffith s Experiment:1928 Frederick Griffith Bacterium - Diplococcus pneumoniae Growth morphology Smooth colonies (capsule): virulent Rough colonies (no capsule): avirulent

4 Conclusion: Something from the dead virulent organisms transformed living cells into virulent cells People spent next 16 years looking at proteins!

5 Experiments Defining the Genetic Material Avery, MacLeod, McCarty:1944 Avery, MacLeod, McCarty- same experiment as Griffith except fractionated virulent cells into various cellular components.

6 Virulence was then transmitted to future generations First of series of experiments that led to the conclusion that DNA was the genetic material.

7 Experiments Defining the Genetic Material Hershey-Chase Experiment: 1952 Hershey-Chase experiment showed that DNA, not protein, is responsible for phage activity in bacterial cells Radioactive phage DNA enters bacteria after attachment, but protein coat of virus remains outside Phage DNA directs the reproduction of virus in infected bacterial cells

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9 Chemical Composition of Nucleic Acids Nucleic acid: organic molecule composed of multiple nucleotides linked in a specific sequence by phosphodiester bonds Nitrogenous base A, G, C, T, U Pentose sugar 5-C sugar (ribose or deoxyribose) Phosphate group (HPO 4 2- )

10 Purines Nitrogenous Bases 9-member ring Adenine (A), guanine (G) Fig. 9-7

11 Nitrogenous Bases Pyrimidines 6-member ring Cytosine (C), Thymine (T), Uracil (U) Fig. 9-7

12 Pentose Sugars RNA & DNA differ by OH at C 2 5 Carbons 5 member ring Fig. 9-7

13 Nucleosides & Nucleotides Fig. 9-8

14 Nucleotide Linkage Assembled 5 3 -PO 42 - on C 5 bonds with OH on C 3 Phosphodiester bond Fig. 9-10

15 DNA

16 Nucleotide Combinations How much variability in sequence is possible with only 4 bases? Ex., nucleotide containing 100 bases = 1.6x10 60 possible nucleotide combinations

17 Watson-Crick Model Deduced structure by synthesizing data from other work on DNA composition & structure G+C composition studies Chargaff X-ray diffraction Astbury; Wilkins & Franklin

18 DNA Base Composition Erwin Chargaff ( ) % A & T always proportional (1:1) % G & C always proportional (1:1) Sum of purines (A+G) = sum of pyrimidines (C+T) G+C typically A+T Table 9.3

19 X-Ray Diffraction Maurice Wilkins & Rosalind Franklin ( ) Purified fibers of DNA and bombarded with X-rays X-rays diffracted based on arrangement of atoms Results suggested... Helical structure 3.4Å (0.34 nm) periodicity Fig. 9-11

20 Watson-Crick Model (1953) Key deductions 2 polynucleotide chains Form R-handed double helix Antiparallel Fig. 9-12

21 Handedness Right Rotates counterclockwise Left Rotates clockwise

22 Watson-Crick Model (1953) Key deductions Nitrogenous bases Flat Perpendicular to axis Stacked 3.4Å apart Located within structure Pair with bases on opposite chain H-bonds G C; A=T Fig. 9-12

23 Watson-Crick Model (1953) Key deductions Helix 20Å diameter Complete turn is 34Å long 10 bases/turn Alternating major & minor grooves appear along axis Fig. 9-12

24 Other Forms of DNA B A normal Aqueous, low salt Dehydrated, high salt C, D, E, P Z All G C Left handed helix Physiological function? Fig. 9-13

25 Useful Properties of DNA Melting temperature (T m ) UV absorption (260 nm) UV absorption with viscosity (unwinding) Use Tm & UV to determine G+C content Fig. 9-14

26 Useful Properties of DNA Hybridization & replication procedures DNA will denature/renature with heat Use to hybridize Known DNA with unknown DNA with known probe Radiolabeled or fluorescent FISH: fluorescence in situ hybridization Use to replicate specific regions of DNA PCR: polymerase chain reaction Purpose Identify homology between DNA sequences Identify specific DNA sequences Produce copies of DNA sequences

27 Useful Properties of DNA Separation by gel electrophoresis Gel creates porous substrate Movement through gel caused by electric field DNA has negative charge Fragments separate by size Smaller molecules move through gel faster Fig. 9-19

28 RNA in contrast to DNA Nitrogenous bases Uracil replaces thymine U binds to A Sugar Ribose replaces deoxyribose Phosphate no change! Single-stranded With some exceptions

29 Major Classes of RNA Ribosomal RNA (rrna) Most abundant Structural component of ribosomes Messenger RNA (mrna) Product of transcription Template for protein synthesis Transfer RNA (trna) Decoder for translation

30 Minor Classes of RNA Small nuclear RNA (snrna) Assists in transcription Telomerase RNA Found at telomeres Assists in DNA synthesis Micro RNA (mirna) Regulates gene transcription Short interfering (sirna) Blocks translation