Organism Organs Tissues Cells (DNA) October 18, DNA is the molecular repository of genetic information. The ability to store and transmit genetic information from one generation to the next is a fundamental condition for life.

October 18, What is the genetic material? Chromosomes are made of protein and DNA. So, one or other or combination of both should be the genetic material. Fundamental property required of the genetic material is that it must be able to exist in an infinite variety of forms: - Each cell contains a large number of different genes - Each gene controls different heritable trait - Each gene should have different structure These speculations led biologists conclude that protein (not DNA) was the genetic material. DNA was believed to be small invariant molecule, whereas proteins were known to be macromolecules made of long polymers of amino acids.

Genetic Material By late 1930sDNA was a long polymer and, like protein, could exist in an almost infinite number of variable forms Identification of the chemical nature of the transforming principle: - Transforming principle can change the bacterium Streptococcus pneumoniae, from one form to another (avirulent to virulent form). - Frederick Griffith1928bacterial transformation Cell membrane Cell wall Slime layer or capsule 3 S = Smooth; with slime layer or capsule; virulent R = Rough; without capsule; avirulent

Griffith’s Experiment (bacterial transformation) IS mouse death IIR mouse alive IS Δ 60 0 C for 3h mouse alive Heat killed IS + living IIR mouse death - Virulent S bacteria can be obtained from the mouse postmortem - Some property of the dead IS bacteria can transform the live IIR into IS - The transforming principle is the genetic material 4

The transforming principle is DNA Oswald Avery, Colin Macleod and Maclyn McCarty1944 Filterate from heat-killed S cells Trypsin, chemotrypsin (degrade protein) SIII enzyme (degrade polysaccharide) Ribonuclease (degrade RNA) Deoxyribonuclease (degrade DNA) transformation No transformation 5

Components of DNA Nucleotides (the monomer in DNA) - the sugar component (pentose; 2’- deoxyribose) - the nitrogenous base (double-ring purines A & G; single-ring pyrimidines T & C) - the phosphoric acid component (three phosphate groups can be attached) 5 carbons in the pentose sugar C1’ base is attached C2’ ribose or deoxyribose C3’ point of attachment for more nucleotides C4’ completes ring via oxygen which bridges to the C1 C5’ hangs away from the ring; point of attachment for its –PO 4 O H HHO H OH HH HOCH 2 1’ 2’3’ 4’ 5’ October 18, 20156

7 Structural elements of the nucleotides October 18, 2015

Polynucleotides: - Nucleotides are joined together by phosphodiester bonds - Polynucleotides have distinct ends (5’-P terminus and 3’-OH terminus) - Polynucleotides can be any length and have any sequence October 18, 20158

9 The double helix J. Watson and F. Crick (Nobel Prize in 1962) 1953 discovered double helix using model building based on all the available information. E. Chargaff demonstrated that the base ratios in DNA are constant. Result: A=T and G=C; total purines (A+G) is equal to total pyrimidines (T+C); but A+T ≠ G+C; GC content varies from species to species

10 1.DNA is double-stranded 2.Sugar-phosphate backbone is on the outside, and the bases are stacked on the inside of the helix 3.Bases of the two strands interact by hydrogen bonding (A-T and G-C) 4.Ten base pairs per turn of the helix; pitch of the helix is 34A 0 i.e. spacing between adjacent base pairs is 3.4A 0 ; the helix is 20A 0 in diameter. 5.The two strands are antiparallel 6.The double helix has two different grooves; major and minor 7.The double helix is right-handed Structure of DNA October 18, 2015 DNA movie

Circumfrance of Earth = 4 X 10 4 Km Sun – Earth distance = 1.5 X 10 8 Km Total length of your DNA = 2 X Km One human being contains ≈ cells One human cell contains 2 meters of DNA October 18,

Supercoiling Relaxed and supercoiled plasmid DNAs October 18,

Bacterial DNA is a compact nucleoid October 18,

Nucleosome is the structural unit of chromatin Chromatin spilling out of lysed nuclei Digestion of chromatin with micrococcal nuclease. Nucleosomes are 10A 0 beads. October 18,

Beads on a String—10 nm Fiber

10 nm filament; nucleosomes histones (= 1g per g DNA) DNA protein purification H1 H3 H2A H2B H4 Basic (arg, lys); + charges bind to - phosphates on DNA

deoxyribonuclease I (DNase I) digestion nucleosomes Conclude: histones in a nucleosome protect 200 nt from DNase I attack. Separate DNA from protein H1 2H3 2H2A 2H2B 2H4 the histone octamer proteins 200 nt fragments DNA

A Nucleosome has 200 bp DNA and core histones bases October 18,

Organization of histone octamer October 18,

Path of nucleosomes in the chromatin fiber 10 nm fiber30 nm fiber October 18, Chromosome movie

Folding of 30 nm fiber Loops of DNA attached to a nuclear scaffold. Numerous loops of DNA attached to a scaffold like structure. October 18,

Net result: Each DNA molecule has been packaged into a chromosome that is 10,000-fold shorter than its extended length Packaging of chromatin fiber Two chromatids (10 coils each) One coil (30 rosettes) One rosette (6 loops) One loop (75000 bp) 30 nm fiber 10 nm fiber DNA October 18,

THANK YOU October 18,