Chapter 16: The Molecular Basis of Inheritance (DNA)

1. Chapter 16: The Molecular Basis of Inheritance (DNA) Zooming in on DNA

2. Mixture of heat-killed S cells and living R cells Heat-killed S cells (control) Living R cells (control) Living S cells (control) LE 16-2 RESULTS Mouse dies Mouse healthy Mouse healthy Mouse dies Living S cells are found in blood sample

3. Phage head LE 16-3 Tail Tail fiber DNA 100 nm Bacterial cell

4. Empty protein shell LE 16-4 Radioactive protein Radioactivity (phage protein) in liquid Phage Bacterial cell Batch 1: Sulfur (35S) DNA Phage DNA Centrifuge Pellet (bacterial cells and contents) Radioactive DNA Batch 2: Phosphorus (32P) Centrifuge Radioactivity (phage DNA) in pellet Pellet Activity: Hersey Chase Experiment

5. Sugar–phosphate backbone Nitrogenous bases 5 end LE 16-5 Thymine (T) Adenine (A) Cytosine (C) Phosphate DNA nucleotide Sugar (deoxyribose) 3 end Guanine (G)

6. DNAi.org Chargaff

7. DNAi.org LE 16-6 Franklin’s X-ray diffraction photograph of DNA Rosalind Franklin

8. DNAi.org Figure 16-01

9. DNAi.org Purine + purine: too wide LE 16-UN298 Pyrimidine + pyrimidine: too narrow Purine + pyrimidine: width consistent with X-ray data

10. LE 16-8 Sugar Sugar Adenine (A) Thymine (T) Sugar Sugar Guanine (G) Cytosine (C)

11. DNAi.org LE 16-7 5 end Hydrogen bond 3 end 1 nm 3.4 nm 3 end 0.34 nm 5 end Key features of DNA structure Partial chemical structure Space-filling model Activity: DNA & RNA Structure Activity: DNA Double Helix

12. LE 16-9_4 The first step in replication is separation of the two DNA strands. Each parental strand now serves as a template that determines the order of nucleotides along a new, complementary strand. The parent molecule has two complementary strands of DNA. Each base is paired by hydrogen bonding with its specific partner, A with T and G with C. The nucleotides are connected to form the sugar-phosphate back- bones of the new strands. Each “daughter” DNA molecule consists of one parental strand and one new strand.

13. Second replication First replication Parent cell Conservative model. The two parental strands reassociate after acting as templates for new strands, thus restoring the parental double helix. LE 16-10 Semiconservative model. The two strands of the parental molecule separate, and each functions as a template for synthesis of a new, comple-mentary strand. Dispersive model. Each strand of both daughter molecules contains a mixture of old and newly synthesized DNA.

14. Bacteria cultured in medium containing 15N Bacteria transferred to medium containing 14N Meselson & Stahl LE 16-11 Less dense DNA sample centrifuged after 20 min (after first replication) DNA sample centrifuged after 40 min (after second replication) More dense Second replication First replication Conservative model Semiconservative model Dispersive model

16. Parental (template) strand 0.25 µm Origin of replication Daughter (new) strand LE 16-12 Replication fork Bubble Two daughter DNA molecules In this micrograph, three replication bubbles are visible along the DNA of a cultured Chinese hamster cell (TEM). In eukaryotes, DNA replication begins at may sites along the giant DNA molecule of each chromosome. Activity: DNA Replication: An Overview

17. New strand Template strand 5¢ end 3¢ end 5¢ end 3¢ end LE 16-13 Sugar Base Phosphate DNA polymerase 3¢ end 3¢ end Pyrophosphate Nucleoside triphosphate 5¢ end 5¢ end Activity: DNA Replication: A Closer Look

18. 3¢ 5¢ Parental DNA Leading strand 5¢ LE 16-14 3¢ Okazaki fragments Lagging strand 3¢ 5¢ DNA pol III Template strand Leading strand Lagging strand Template strand DNA ligase Overall direction of replication

19. Primase joins RNA nucleotides into a primer. 3¢ 5¢ 3¢ 5¢ Template strand DNA pol III adds DNA nucleotides to the primer, forming an Okazaki fragment. LE 16-15_6 3¢ 5¢ RNA primer 3¢ 5¢ After reaching the next RNA primer (not shown), DNA pol III falls off. Okazaki fragment 3¢ 3¢ 5¢ 5¢ After the second fragment is primed, DNA pol III adds DNA nucleotides until it reaches the first primer and falls off. 5¢ 3¢ 3¢ 5¢ DNA pol I replaces the RNA with DNA, adding to the 3¢ end of fragment 2. 5¢ 3¢ 3¢ 5¢ DNA ligase forms a bond between the newest DNA and the adjacent DNA of fragment 1. The lagging strand in the region is now complete. 5¢ 3¢ 3¢ 5¢ Overall direction of replication

20. DNAi.org LE 16-16 Overall direction of replication Lagging strand Leading strand Origin of replication Leading strand Lagging strand OVERVIEW DNA pol III Leading strand DNA ligase Replication fork 5¢ DNA pol I 3¢ Primase Lagging strand Parental DNA DNA pol III Primer 3¢ 5¢ Activity: DNA Replication: A Review

21. A thymine dimer distorts the DNA molecule. Proofreading and Repair LE 16-17 A nuclease enzyme cuts the damaged DNA strand at two points and the damaged section is removed. Nuclease Repair synthesis by a DNA polymerase fills in the missing nucleotides. DNA polymerase DNA ligase DNA ligase seals the free end of the new DNA to the old DNA, making the strand complete.

22. 5¢ Leading strand End of parental DNA strands Lagging strand 3¢ LE 16-18 Last fragment Previous fragment RNA primer Telomeres Lagging strand 5¢ 3¢ Primer removed but cannot be replaced with DNA because no 3¢ end available for DNA polymerase Removal of primers and replacement with DNA where a 3¢ end is available 5¢ 3¢ Second round of replication 5¢ New leading strand 3¢ New leading strand 5¢ 3¢ Further rounds of replication Shorter and shorter daughter molecules