DNA STRUCTURE AND ANALYSIS I.

CHARACTERISTICS OF THE GENETIC MATERIAL Replication Storage of Information Expression of Information Variation by mutation

II. PROTEIN AS THE GENETIC MATERIAL (until 1944) Chromosomes have a nucleic acid and a protein component. Both components were candidates for the role of genetic material

Three factors that favoured protein as the genetic material 1.Abundance of protein in cells Account for 50% of the dry weight of cells 2.Accepted proposal for the chemical structure of nucleic acids Tetranucleotide Structure of DNA by Phoebus Levene 3.Areas of most active research in genetics Transmission genetics and mutation

TETRANUCLEOTIDE HYPOTHESIS (1910)

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 1

Nucleic acids were thought to be simple repetitive polymers

o Simple 4-nucleotide unit repeated over and over in DNA III. EVIDENCE FAVORING DNA AS THE GENETIC MATERIAL

Frederick Griffiths Transformation Experiment (1927)

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 2

Transforming Principle: The Avery, MacLeod, and McCarty Experiment (1944) o Oswald Theodore Avery (1877 1955) o Colin Munro MacLeod (1909 1972) o Maclyn McCarty (1911 - 2005)

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 3

The molecule responsible for transformation was DNA (deoxyribonucleic acid).

The Hershey-Chase Experiment (1952) o Alfred Hershey (1908 1997) o Martha Chase (1927 2003)

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 4

Study of bacterium E.coli and one of its infecting viruses, bacteriophage T2

T2 phages consist of approximately 50% protein and 50% DNA. Infection is initiated by adsorption of the phage by its tail fibers to the bacterial cell wall. The production of new viruses occurs within the bacterial cell.

They used radioisotopes 32P and 35S to follow the molecular components of phages during infection.
o
32

P effectively labels DNA DNA contains phosphorus

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 5

35

S effectively labels protein Protein contains sulfur

IV. NUCLEIC ACID CHEMISTRY A. Nucleotides o Building blocks of nucleic acids o Components:

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 6

 Nitrogenous base Purine (nine-member double ring) Pyrimidine (six-member single ring) Pentose sugar Ribose Deoxyribose Phosphate groups

B. Nucleosides o Composed of nitrogenous base and pentose sugar

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 7

 The bonding among the nucleotides is highly specific. C-1 atom of sugar links with the nitrogenous base. o Purine (N-9 atom) o Pyrimidine (N-1 atom) C-2, C-3 and C-5 atom of sugar links with the phosphate groups o C-5 phosphate configuration is the prevalent form in biological systems; one found in DNA and RNA

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 8

V. DNA STRUCTURE AND ITS FUNCTION A. Base Composition Studies Erwin Chargaff o Used chromatographic methods to separate the four nitrogenous bases in the DNA samples from various organisms o Used quantitative methods to determine the amounts of the four bases from each source

The amount of adenine residues is proportional to the amount of thymine residues in the DNA of any species. The amount of guanine residues is proportional to the amount of cytosine residues. (A+G) = (C+T)

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 9

 The percentage of C+G does not necessarily equal the percentage of A+T B.

X-ray Diffraction Analysis The pattern of scatter (diffraction) can be captured as spots on photographic film.

William Astbury o (1947) periodicity within the structure of the molecule of 3.4 o Bases stacked like coins

Rosalind Franklin o Obtained improved X-ray data from purified samples of DNA o Confirmed the 3.4 periodicity o Suggested that DNA structure is a helix

Linus Pauling o Used diffraction analysis in the study of protein structure

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 10

o Analyzed the work of Astbury and others o Proposed that DNA is a triple helix C. Watson and Crick Model

Two primary sources crucial to the development of James Watson and Francis Cricks model: 1.Base composition studies of samples of DNA 2.X-ray diffraction studies of DNA Watson-Crick Double Helix

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 11

1.

Two long polynucleotide chains are coiled around a central axis, forming a right-handed double helix. The two chains are antiparallel. C-5-to-C-3 orientations run in opposite directions The bases of both chains are flat structures, lying perpendicular to the axis. stacked on one another 3.4 (0.34 nm) apart

2.

3.

4.Hydrogen bonding A pairs with T (double bond) G pairs with C (triple bond) 5.Each complete turn of the helix 34 (3.4 nm) long 10 bases per turn in each chain
6.

Alternating larger major grooves and smaller minor grooves

7.Diameter of helix = 20 (2.0 nm) Accurate Analysis of the form of DNA

Structure No. of bases per turn Each base pair rotation around the helical axis relative to the adjacent base pair Recent, more accurate 10.4 34.6 Watson-Crick Model 10.0 36

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 12

Base pairs per turn

More than 10 bp

10 bp

VI. ALTERNATIVE FORMS OF DNA

Characteri stics Condition Base pairs per turn Diameter Configurati on A-DNA High salt or dehydrati on 9 bp 23 Righthanded Tilted and displaced laterally IRT the helical axis B-DNA Low salt condition 10.4 bp 20 Righthanded C-DNA Greater dehydrati on (lab) 9.3 bp 19 -DE-DNA DNA Helices Helices lacking lacking guanin guanin e e 8 bp --7 bp --Z-DNA PDNA -2.62 bp ---

-12 bp 18 Lefthanded Zigzag conformati on (major groove present in B-DNA nearly eliminated )

Arrangeme nt of base pairs

Lying flat; Perpendicu lar to the helical axis

Same with ADNA

--

--

--

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 13

Klug, W., Cummings, M., and Spencer, C. 2006. Concepts of Genetics. 8th ed. Pearson Prentice Hall. Page 14