1. Chapter 16 P. 305 - 324

3. 16.1 Dna Is The Genetic Material T.H Morgan’s group: showed that genes are located along chromosomes. Two chemical components of chromosomes are DNA and protein. Little was known about nucleic acids. Role of DNA in heredity was first worked out by studying bacteria and the viruses that infect them.

4. Important Scientists in the Discovery of DNA Frederick Griffith Oswald Avery Alfred Hershey and Martha Chase Rosalind Franklin Francis Crick and James Watson

5. Frederick Griffith Discovery of role in 1928 Vaccine against pneumonia (mice) Frederick Griffith studied Streptococcus pneumoniae Two stains of the bacterium Pathogenic Non pathogenic Heated the pathogenic and killed the bacteria. Mixed the cell remains with living bacteria of the nonpathogenic and found some cells were then pathogenic

6. Frederick Griffith This newly acquired trait was inherited by all the descendants of the transformed bacteria. Called the phenomenon transformation: a change in genotype and phenotype due to the assimilation of external DNA by a cell.

7. Oswald Avery Identity of transforming substance Three main candidates DNA RNA Protein Avery broke open the heat-killed bacteria and extracted the cellular contents Special treatments to inactivate each of the three molecules

8. Oswald Avery Tested each for its ability to transform live nonpathogenic bacteria. DNA was left active – transformation occurred Transforming agent was then announced as DNA Studied viruses for more information Bacteriophages (phages): bacteria-eaters A virus is composed of DNA(or RNA) enclosed by a protective coat.

9. Hershey and Chase Devised an experiment showing that only one of the two components enters the E.coli cell. Specifically looked at T2 T2 invades Escherichia coli bacteria Radioactive isotope of sulfur to tag protein, and phosphorus to tag DNA.

10. Fig. 16-4-3 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. Rosalind Franklin Used X-Ray crystallography to find out structure of DNA molecules X near center shows DNA twists around Angle of the X suggests two strands and the nitrogenous bases are near the center of the molecule Shows diameter of the double helix

12. Francis Crick and James Watson Built three-dimensional models of DNA Used Rosalind Franklin’s x-ray pictures of DNA to assist in the model The Double Helix Width suggested that it was made up of two strands. Began to build models that would conform to the X-ray measurements and the chemistry of DNA.

13. Watson and Crick- Double Helix Composed of two complementary strands of DNA wrapped around each other Uniform diameter Hydrogen bonds held the two strands together Two hydrogen bonds between A and T Three hydrogen bonds between C and G.

14. Chargaff’s Rule Studied percentages of nitrogenous bases. Almost equal %’s Adenine bonds to Thymine – Guanine bonds to Cytosine = support Nitrogenous Bases make up DNA molecules The two types are: Purines Two rings in the structure Pyrimidines One ring in the structure

15. Chargaff’s Rule

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

18. Base Pairing Watson and Crick stated their hypothesis Pair of templates, each of which is complementary to the other. Prior to duplication, the hydrogen bonds are broken The two chains unwind and separate Each chain acts as a template Eventually, two pairs of chains will result.

19. DNA Replication In DNA replication, the parent molecule unwinds, and two new daughter strands are built based on base- pairing rules When a cell copies a DNA molecule, each strand serves as a template for ordering nucleotides into a new, complementary strand. Nucleotides line up along the template strand and are linked Where there was one double-stranded DNA molecule at the beginning, there are then two at the end. The copying mechanism is analogous to using a photographic negative to make a positive

20. Watson and Crick’s Hypothesis Figure 16.9 A T G C TA TA G C (a) Parent molecule AT GC T A T A GC (c) “Daughter” DNA molecules, each consisting of one parental strand and one new strand (b) Separation of strands A T G C TA TA G C A T G C T A T A G C

21. Replication Models Remained untested for many years Difficult to perform Watson and Crick predicted the semiconservative model Each daughter molecule will have one old strand (derived or “conserved” from the parent molecule) and one newly made strand There are two others: Conservative Dispersive

22. Fig. 16-10 Parent cell First replication Second replication (a) Conservative model (b) Semiconserva- tive model (c) Dispersive model

23. DNA Replication Models Conservative The two parental strands reassociate after acting as templates for new strands. Semiconservative The two strands of the parental molecule separate, and each functions as a template for synthesis of a new, complementary strand. Dispersive Each strand of both daughter molecules contains a mixture of old and newly synthesized DNA.

24. Semiconservative Model 1950 Matthew Meselson and Franklin Stahl devised a clever experiment that supported the semiconservative model. Widely acknowledged among biologists to be a classic example of elegant experimental design. Figure 16.11 shows the experiment performed by Meselson and Stahl.

25. DNA and Replication in Prokaryotes Prokaryotes: ring of chromosome holds nearly all of the cell’s genetic material DNA replication begins at a single point and continues to replicate whole circular strand Replication goes in both directions around the DNA (begins with replication fork)

26. Prokaryotes Eukaryotes

27. Eukaryotic DNA Replication The replication of a DNA molecule begins at special sites called origins of replication Begins in hundreds of locations along the chromosome Begins when the DNA molecule “unzips” creating: Replication fork Replication “bubble”

28. Eukaryotic DNA Replication Hydrogen bonds between base pairs breaks Helicases – enzymes that untwist the double helix at the replication forks. Single-strand binding proteins bind to the unpaired DNA strands, stabilizing them. Topoisomerase – relieves pressure of DNA ahead of replication fork RNA Primer – RNA chain Primase – enzyme that synthesizes the primer

29. Helicase will start to unwind the DNA strand. Topoisomerase will hold the strands together and prevent breaking Single-stranded binding proteins will stabilize the DNA strands The Primase will start to form the RNA chain

30. Synthesizing a New DNA Strand DNA polymerase: catalyze the synthesis of new DNA by adding nucleotides to a preexisting chain. E. coli – DNA polymerase III and DNA polymerase I Humans – 11 different DNA polymerase molecules Most DNA polymerases require a primer and a DNA template strand. DNA polymerase III adds a DNA nucleotide to the RNA primer and then continues adding DNA nucleotides complementary to the parent DNA template strand.

31. Antiparallel Elongation The two strands of DNA in a double helix are antiparallel (0riented in opposite directions). DNA polymerases can only add nucleotides to the free 3’ end of a primer or growing strand. NEVER THE 5’ A new strand can only elongate in the 5’-3’ direction ALWAYS Read in the 3’ – 5’ direction Created in 5’ – 3’ direction

32. Antiparallel Elongation Leading Strand –only 1 primer needed, moves toward the replication fork Lagging Strand – many primers needed, moves away from the replication fork Okazaki Fragments – on lagging strand, short segment of DNA synthesized away from the replication fork DNA ligase – enzyme, joins the sugar-phosphate backbones of all the Okazaki fragments into a continuous DNA strand

34. The DNA Replication Complex By interacting with other proteins at the fork, primase acts as a molecular brake, slowing progress of the replication fork. The DNA replication complex does not move along the DNA The DNA moves through the complex Recent studies support: Two DNA polymerase molecules “reel in” the parental DNA The lagging strand is looped back through the complex Enables more Okazaki fragments to be synthesized in less time

36. Proofreading and Repairing DNA During DNA replication, DNA polymerases proofread each nucleotide against its template as soon as it is added to the growing strand. The polymerase removed the incorrectly paired nucleotide and resumes synthesis. Mismatched nucleotides sometimes are missed. Can also arise after replication Mismatched repair – enzymes remove and replace incorrectly paired nucleotides that have resulted from replication errors.

37. Proofreading and Repairing DNA Most cellular systems that repair incorrectly paired nucleotides use a mechanism that takes advantage of the base-paired structure of DNA. Nuclease – DNA-cutting enzyme. Cuts out the segment of the strand containing the damaged segment. Enzymes involved in filling gaps: DNA polymerase and DNA ligase Nucleotide excision repair – repair system, Figure 16.18

38. Replicating the Ends of DNA Telomeres Found at the ends of each chromosome and contain no genes Can be cut short will not affect normal functioning TTAGGG is the six-nucleotide sequence Telomerase lengthens telomeres in gametes The shortening of telomeres might protect cells from cancerous growth by limiting the number of cell divisions There is evidence of telomerase activity in cancer cells, which may allow cancer cells to persist 

39. Important Enzymes to Remember Helicase, single-strand binding protein, topoisomerase Primase Synthesis of RNA primer DNA polymerase III (DNA pol III) Add new bases to DNA strand DNA polymerase I (DNA pol I) Removes and replaces RNA primer from 5’ end DNA ligase Links Okazaki fragments and replaces RNA primer from 3’ end

41. Vocab Nucleoid –A dense region of DNA in a prokaryotic cell Chromatin –complex of DNA and proteins that makes up a eukaryotic chromosome Heterochromatin – Eukaryotic chromatin that remains highly compacted during interphase and is generally not transcribed. Euchromatin – The less condensed form of eukaryotic chromatin that is available for transcription.

42. Chromosome Structures Bacteria: one double-stranded, circular DNA molecule that is associated with a small amount of protein. Prokaryotes: Ring of chromosomes Holds nearly all the cell’s genetic material Eukaryotes: DNA in chromosomes Found in nucleus

43. Chromatin Packing In the cell, eukaryotic DNA is combined with large amounts of protien. Complex of DNA and protein – chromatin Histones - proteins that are responsible for the first level of DNA packing in chromatin Form a tight bond because DNA is negatively charged and the histones have a positive charge

44. Fig. 16-21a DNA double helix (2 nm in diameter) Nucleosome (10 nm in diameter) Histones Histone tail H1 DNA, the double helixHistones Nucleosomes, or “beads on a string” (10-nm fiber)

45. Fig. 16-21b 30-nm fiber Chromatid (700 nm) LoopsScaffold 300-nm fiber Replicated chromosome (1,400 nm) 30-nm fiber Looped domains (300-nm fiber) Metaphase chromosome

46. Chromosome Organizations 10-nm fiber 30 – nm fiber 300-nm fiber

47. 10 - nm fiber DNA winds around histones to form nucleosome beads Nucleosomes are strung together The string between the beads is called linker DNA Nucleosome consists of DNA wound twice around a protein core composed of two molecules each.

48. 30-nm fiber 10-nm coils Forms a chromatin fiber 30 nm thick Interactions between nucleosomes cause the thin fiber to coil or fold into this thicker fiber

49. 300-nm Fiber 30 nm fiber forms loops called looped domains attached to a chromosome scaffold made of proteins Scaffold is rich in one type of topoisomerase.

50. Heterochromatin and Euchromatin Heterochromatin During interphase, a few regions of chromatin are highly condensed into heterochromatin Dense packing of the heterochromatin makes it difficult for the cell to express genetic information coded in these regions Euchromatin Most chromatin is loosely packed in the nucleus during interphase Condenses prior to mitosis

51. Questions Short answer Using a Venn diagram, compare and contrast prokaryotic and eukaryotic DNA and DNA replication. Include at least 2 similarities and 2 unique characteristics for each cell type.

52. Questions 1. _______ removes section of DNA that is damaged 2. _______ proofreads and repairs damaged/mismatched DNA; base pairing 3. _______ synthesis of RNA primer 4. _______ Links Okazaki fragments; replaces RNA primer from 3’ end (in both leading and lagging strand). 5. _______ relieves pressure of DNA ahead of replication fork 6. _______ attach to separated DNA strands to ensure they stay separated 7. _______ breaks hydrogen bonds between DNA strands 8. _______ lengthens telomeres in gametes DNA ligase, DNA polymerase, Helicase, Primase, Telomerase Single-strand binding proteins, Topoisomerase, Nuclease