How Do Genes Work?

1. How Do Genes Work?

2. Are Genes Composed of DNA or Protein? • Chromosomes, the known carriers of genes, are made of DNA and proteins • Proteins are more variable in structure than DNA, whichwas not thought to be sufficiently complex to code for allthe operations of a cell. • DNA = a polymer made of 4 different types of nucleotides (adenine, cytosine, guanine, thymine) • Protein = a polymer made of 20 different types of amino acids

3. Experiments showing DNA is the genetic material SMOOTH COLONY (S) ROUGH COLONY (R) There are two strains of Streptococcus pneumoniae. R strain is benign (Lacks a protective capsule, recognized and destroyed by host’s immune system) S strain is virulent (Polysaccharide capsule prevents detection by host’s immune system)

4. Experiments by Griffiths, 1928 Conclusion:

5. Experiments by Avery, MacLeod and McCarty (1944) 1. Remove lipids and carbohydrates from a solution of heat-killed S cells. Proteins, RNA, and DNA remain. Heat-killed S cells Lipids Carbohydrates Add proteinases Add DNAse Add RNAse Add deoxy- ribonuclease Add proteinases Add ribonuclease 2. Treat solutions with enzymes to destroy proteins, RNA, or DNA. Sample should contain NO DNA Sample should contain NO PROTEIN Sample should contain NO RNA Sample should Contain NO PROTEIN Sample should Contain NO RNA Sample should Contain NO DNA 3. Add a portion of each sample to a culture containing R cells. Observe if transformation occurs. Add R cells Add R cells Add R cells S cells appear S cells appear No S cells appear Transformation occurs No transformation occurs Conclusion: Transformation cannot occur unless DNA is present. Therefore, DNA must be the hereditary material.

6. Are Genes Composed of DNA or Protein? • Transformation experiments provide the first evidence that genes are DNA. • Griffith: material from dead virulent Streptococcus can transform benign Streptococcus into a virulent strain. • Avery et al.: extracted material from dead virulent Streptococcus and treated it with enzymes to destroy either DNA or RNAor proteins. Only DNA-destroying enzymes prevented transformation, hence DNA is the genetic material.

7. Are Genes Composed of DNA or Protein? • Not everyone was convinced • Hershey and Chase (1952): T2 virus experiments convinces skeptics that DNA is the genetic material since the material injected by the virus into host cells is DNA, not protein.

8. Bacteriophages are viruses that infect bacterial cells

9. How do viruses work? Virus particle Virus particle Host cell genome Host cell genome 1. Viral genome enters host cell. 1. Viral genome enters host cell. DNA mRNA 2. Viral genome is replicated and transcribed. Virus particle Host cell genome New viruses released 1. Viral genome enters host cell. DNA mRNA 4. Particles assembled inside host. 2. Viral genome is replicated and transcribed. Protein 3. Viral particles produced

10. Experiments by Hershey and Chase (1952) • Protein contains sulfur, but not phosphorous • DNA contains phosphorous, but not sulfur • Growing viruses with radioactive sulfur will label proteins but not DNA • Growing viruses with radioactive phosphorous will label DNA but not proteins

11. Experiments by Hershey and Chase (1952)

12. Sister chromatids DNA replication Centromere One chromosome (Unreplicated state) One chromosome (Replicated state) The structure and replication of genetic material • DNA structure was determined by James Watson and Francis Crick using data from Rosalind Franklin • In order to pass on genetic material from parent (cell) to offspring (cell), the genetic material must be duplicated: DNA replication

13. Figure 16.0 Watson and Crick

14. Figure 16.4 Rosalind Franklin and her X-ray diffraction photo of DNA

15. Free DNA nucleotides are called dNTPs P Structure of dNTPs P P CH2 5' Base O 4' 1' 2' 3' OH

16. Nucleotide Pyrimidines NH2 O O H3C NH O N NH N 5 –O Nitrogenous base P O O N N N O O O O– 1 4 Phosphate group H H H 2 3 Cytosine (C) Uracil (U) Thymine (T) 5-carbon sugar Purines NH2 O Deoxyribose Ribose N N NH N OH HO5CH2 OH HO5CH2 O O H H H H 4C 1C 4C 1C N N N N NH2 C3 2C C3 2C H H H H H H OH OH OH H Adenine (A) Guanine (G) Review of DNA structure: DNA and RNA are polymers made of nucleotide subunits

17. A single strand of nucleotides is made when a phosphodiester bond is formed between the 3’ C of one nucleotide and the 5’ C of another 5' 3'

18. 5´ 3´ 5´ 3´ 5´ 3´ T T A A DNA is a double helix. C G G C G C C G A A T T T A T A C G C G G C G A T A T A T T A C G C G T A A T G C T A A T G G C C A T T A 5´ 3´ 3´ 5´ 5´ 3´ Cartoon of base pairing Cartoon of double helix Space-filling model of double helix DNA is double stranded with the 2 strands held together by hydrogen bonds between complementary bases

19. Figure 16.6 Base pairing in DNA

20. Unnumbered Figure (page 292) Purine and pyridimine

21. Distinguishing between Models of DNA Replication • Three different models of how DNA might replicate were proposed based on DNA structure. • Semi-conservative replication • Conservative replication • Dispersive replication

22. Figure 16.8 Three alternative models of DNA replication

23. Distinguishing Between Models of DNA Replication • The Meselsohn and Stahl experiment determines which model is correct. • 15N was fed to growing E. coli cells to mark DNA (“heavy” DNA), then cells were switched to 14N. • Newly synthesized DNA will incorporate 14N (“light” DNA. 14N 15N 15N 15N 15N 14N

24. Figure 16.9 The Meselson-Stahl experiment tested three models of DNA replication (Layer 1)

25. Figure 16.9 The Meselson-Stahl experiment tested three models of DNA replication (Layer 2)

26. Figure 16.9 The Meselson-Stahl experiment tested three models of DNA replication (Layer 3)

27. Figure 16.9 The Meselson-Stahl experiment tested three models of DNA replication (Layer 4)