1. Chapter 16 Molecular Basis of Inheritance (DNA structure and Replication) Helicase Enzyme

2. What is the genetic material? DNA or protein? The Amazing Race

3. 1928 Griffith – transformation of pneumonia bacterium

4. 1944 Avery – further studied transformation by destroying lipids, CHO, and proteins

5. 1947 Chargaff – Quantified purines and pyrimidines Suggested base pairing rules (A=T, C=G)

6. 1950 Wilkins and Franklin – DNA X-rays (a) Rosalind Franklin (b) Franklin’s X-ray diffraction photograph of DNA

7. 1952 Hershey and Chase – bacteriophages – incorporation of radioactive viral DNA in new phages

8. EXPERIMENT Phage DNA Bacterial cell Radioactive protein Radioactive DNA Batch 1: radioactive sulfur ( 35 S) Batch 2: radioactive phosphorus ( 32 P)

9. EXPERIMENT Phage DNA Bacterial cell Radioactive protein Radioactive DNA Batch 1: radioactive sulfur ( 35 S) Batch 2: radioactive phosphorus ( 32 P) Empty protein shell Phage DNA

10. EXPERIMENT Phage DNA Bacterial cell Radioactive protein Radioactive DNA Batch 1: radioactive sulfur ( 35 S) Batch 2: radioactive phosphorus ( 32 P) Empty protein shell Phage DNA Centrifuge Pellet Pellet (bacterial cells and contents) Radioactivity (phage protein) in liquid Radioactivity (phage DNA) in pellet

11. 1953 Watson and Crick – DNA Model

12. 1962 Nobel Prize awarded to Watson and Crick and Wilkins ** Conclusion: DNA = Genetic Material, not Protein

13. Models of DNA Replication

14. Semi-Conservative Model (1950s - Meselson and Stahl)

15. Fun DNA Replication Facts 6 billion bases in human cell = 2 hours of replication time 500 nucleotides added per second Accurate (errors only 1 in 10,000 base pairs)

16. Anti- Parallel Structure of DNA

17. Mechanism of Replication Step 1 Origins of Replication = Special site(s) on DNA w/Specific sequence of nucleotides where replication beginsOrigins of Replication –Prokaryotic Cells = 1 site (circular DNA) –Eukaryotic Cells = several sites (strands)

18. Steps 2 - 5 Helicase: (enzyme) unwinds DNA helix forming a “Y” shaped replication fork on DNA Replication occurs in two directions, forming a replication bubble To keep strands separate, DNA binding proteins attach to each strand of DNA Topoisomerases: enzymes that work w/helicase to prevent “knots” during unwinding.

19. Step 6 - Priming Priming = due to physical limitation of DNA Polymerase, which can only add DNA nucleotides to an existing chain RNA primase – initiates DNA replication at Origin of Replication by adding short segments of RNA nucleotides. Later these RNA segments are replaced by DNA nucleotides by DNA Pol.

20. Step 7 DNA Pol. = enzyme that elongates new DNA strand by adding proper nucleotides that base- pair with parental DNA template DNA Pol. can only add nucleotides to the 3’ end of new DNA, so replication occurs from a 5’ to 3’ direction Leading vs. Lagging Strand results

21. LeadingLeading vs. Lagging StrandLagging Strand Leading Strand: strand that can elongate continuously as the replication for progresses Lagging Strand: strand that cannot elongate continuously and moves away from replication fork. Short Okazaki fragments are added from a 5’ to 3’ direction, as replication fork progresses.

22. 3’ 5’ 3’5’ 3’ 5’

23. Step 8 DNA Ligase = enzyme that “ligates” or covalently bonds the Sugar-Phosphate backbone of the short Okazaki fragments together Primers are required prior to EACH Okazaki fragment

24. DNA i Flash Overview

25. Step 10: Fixing Errors DNA Pol. Proofreads as it elongates Special enzymes fix a mismatch nucleotide pairs Excision Repair: –Nuclease: Enzyme that cuts damaged segment –DNA Pol. Fills in gap with new nucleotide

26. Mutations Thymine Dimers (covalent bonding btwn Thymine bases) –often caused by over- exposure to UV rays  DNA buckeling  skin cancer results, unless corrected by excision repair Substitutions: incorrect pairing of nucleotides Insertions and Deletions: an extra or missing nucleotide  causes “frameshift” mutations (when nucleotides are displaced one position)

27. Problems with Replication Since DNA Polymerase can only add to a 3’ end of a growing chain, the gap from the initial 5’ end can not be filled Therefore DNA gets shorter and shorter after each round of replication

28. Solution? Bacteria have circular DNA (not a problem) Ends of some eukaryotic chromosomes have telomeres at the ends (repeating nucleotide sequence that do not code for any genes) Telomeres can get shorter w/o compromising genes Telomerase = enzyme that elongates telomeres since telomeres will shorten

29. Telomerases are not in most organisms Most multicellular organisms do not have telomerases that elongate telomeres (humans don’t have them) So, telomeres = limiting factor in life span of certain tissues Older individuals typically have shorter telomeres