Ch. 13 - DNA - Structure & Function

1. Ch. 13 - DNA - Structure & Function DNA Structure & Function

2. DNA as Genetic Material • Johann Miescher (1869) • Removed nuclei from pus cells • Found they contained a chemical he called nuclein • This was rich in phosphorus and had no sulfur; thus it could not be a protein Later scientists realized there were two types of nucleic acids: DNA (deoxyribonucleic acid) and RNA (ribonucleic acid)

3. Frederick Griffith (1931) • Investigated virulence of Streptococcus pneumoniae in mice in following manner: • 1. S strain bacteria have a smooth capsule & are capable of killing mice • 2. R strain have no capsule & don’t kill mice • 3. Injected heat-killed S strain bacteria into mice; they did not die • 4. Injected mice with mixture of heat-killed S strain & live R strain. These mice had living S strain bacteria & died • Concluded that virulence passed from the dead strain to the living strain; transformation had occurred

4. Griffith’s Transformation Experiment

5. Avery, MacLeod & McCarty (1944) • (Refer to transparency here first) • Discovered that DNA is the transforming substance. 1. Took DNA only from the S bacteria and mixed it with R bacteria. 2. S strain DNA was then incorporated into genome of living R strain bacteria and they were then transformed into S strain bacteria. 3. Enzymes that degrade proteins or RNA did not prevent transformation while those that digest DNA did.

6. Reproduction of Viruses • Viruses consist of a protein coat (capsid) surrounding a nucleic acid core • Bacteriophages are viruses that infect bacteria

7. Hershey and Chase (1952) • Did an experiment to determine whether the bacteriophages inject the protein or DNA into the bacteria. • Radioactively labeled the DNA core and protein capsid of a bacteriophage 1. Radioactive P (found in DNA & not in protein) was found inside cells 2. Radioactive S (found in protein & not in DNA) was found mainly outside of cells • Results indicated that DNA, not the protein, enters the host • The DNA of the phage contains genetic information for producing new phages

8. Hershey and Chase Experiments

9. Structure of DNA • DNA contains: • Two nucleotides with purine bases. These are double ring nitrogenous bases. • Adenine (A) • Guanine (G) • Two nucleotides with pyrimidine bases. These are single ring nitrogenous bases. • Thymine (T) • Cytosine (C)

10. Nucleotide Composition of DNA

11. Chargaff’s Rules • The amounts of A, T, G, and C in DNA: • Identical in identical twins • Varies between individuals of a species • Varies more from species to species • In each species, there are equal amounts of: • A & T • G & C • All this suggests DNA uses complementary base pairing to store genetic information. • Human chromosome estimated to contain, on average, 140 million base pairs. • Number of possible nucleotide sequences 4^140,000,000.

12. Diffraction Data • Rosalind Franklin: • Studied structure of DNA using X-rays. • Found that if a concentrated solution of DNA is made it forms into a crystal like structure. • When X-rayed, an X-ray diffraction pattern results. • The pattern of DNA shows that it is a helix.

13. X-Ray Diffraction of DNA

14. Watson and Crick Model (1953) • Using data provided by Franklin’s X-ray diffraction and other knowledge about DNA, they eventually determined that DNA is a double-helix • Sugar-phosphate backbones make up the sides • Hydrogen-bonded bases make up the rungs. Complementary bases (A-T; C-G) pair up. • Model matched data of both Franklin & Chargaff • Received a Nobel Prize in 1962

15. Watson/Crick Model of DNA

16. DNA Replication: • Replication = process of copying a DNA molecule • 1. During DNA replication, each old DNA strand of the parental molecule (original double helix) serves as a template for a new strand in a daughter molecule. • 2. DNA replication is termed semiconservative replication because one of the old strands is conserved, or present, in each daughter DNA molecule.

17. Steps of Replication • 1. Unwinding • DNA replication begins at numerous points along linear chromosome called replication forks. • DNA unwinds and unzips into two strands. Weak hydrogen bonds between paired bases are broken. • A special enzyme, DNA helicase, unwinds the DNA.

18. Replication (cont’d) 2. Complementary base pairing • Each old strand of DNA serves as a template for a new strand • New complementary nucleotides are positioned by process of complementary base pairing • A special enzyme, called DNA polymerase, helps to position the complementary base pairs

19. Replication (cont’d) • 3. Joining • The complementary nucleotides join to form new strands. • This is also helped by DNA polymerase

20. Semiconservative Replicationof DNA

21. Meselson & Stahl’s experiment (1958) • Confirmed semiconservative replication theory • They grew bacteria in a medium containing heavy N-15 so only heavy DNAs were found. • Switched bacteria to N-14 medium. • After 1 division, only hybrid DNA was found • After 2 divisions, half the DNA is light & half is hybrid • These are the results expected if DNA replication is semiconservative.

22. Meselson and Stahl’sDNA replication experiment

23. Details of DNA Replication • Carbon atoms are numbered in the deoxyribose molecule. • DNA strands are antiparallel. One of the strands runs from 3’ to 5’ in one direction, and the other strand runs from 3’ to 5’ in the opposite direction. • During replication, DNA polymerase has to synthesize the daughter strand in the 5’ to 3’ direction. • Why? DNA polymerase can only join a nucleotide to a free 3’ end of a previous nucleotide.

25. Details of DNA Replication (cont’d) • This also means that DNA polymerase cannot start the synthesis of a DNA chain. • An RNA polymerase lays down a short amount of RNA, called an RNA primer,that is complementary to DNA. • Then DNA polymerase can join DNA nucleotides to the 3’ end of the growing daughter strand.

26. Details of DNA Replication (cont’d) • As helicase unwinds DNA, one parental strand runs in the 3’ to 5’ direction toward the fork. Thus, the new complementary daughter strand will be synthesized from the 5’ to 3’ direction. This strand is called the leading strand. • The other parental strand, however, is running in the opposite direction (3’ to 5’ AWAY from the fork). The daughter strand must begin at the fork and run in the opposite direction to the leading strand. This is called the lagging strand.

27. Antiparallel Replication of DNA

28. Details of DNA Replication (cont’d) • Replication of the lagging strand is discontinuous. • It results in segments called Okazaki fragments. • While proofreading, DNA polymerase will remove the RNA primers and replace them with complementary DNA nucleotides. • DNA ligase will then join the fragments together.

29. Antiparallel Replication of DNA

30. DNA Replication:Prokaryotic • Prokaryotic Replication • Bacteria have a single circular loop • Replication moves around the circular DNA molecule in both directions. Takes about 40 minutes. • Produces two identical circles • Cell divides between circles, as fast as every 20 minutes

31. Replication:Prokaryotic vs. Eukaryotic

32. Replication Errors • Genetic variations are the raw material for evolutionary change • Mutation: • A permanent (but unplanned) change in base-pair sequence • Some due to errors in DNA replication. Proofreading occurs which eliminates most errors. Mistake rate is only 1 per 1 billion base pairs. • Others are due to DNA damage like UV radiation • DNA repair enzymes are usually available to reverse most errors

33. Videos for Chapter 13 DNA Replication Animation I http://www.courses.fas.harvard.edu/~biotext/animations/replication1.html DNA Replication Animation II http://highered.mcgraw-hill.com/olc/dl/120076/bio23.swf