SPU-22: The Unity of Science from the Big Bang to the Brontosaurus and Beyond

1. SPU-22: The Unity of Science from the Big Bang to the Brontosaurus and Beyond Lecture 20 16 April 2014 Science Center Lecture Hall A

2. Today’s Topics Hershey-Chase experiment: How DNA decisively chosen over protein Watson-Crick model of DNA: How developed (demonstration of result) Meselson-Stahl experiment: How DNA replicates

3. Determined Detectives: I Alfred Hershey and Martha Chase Q: What is responsible for heredity – protein or DNA (despite Avery)? A: Use radioactive tracers, 32P (DNA) and 35S (protein), in virus (= DNA wrapped in protein) that infects benign bacteria – no phosphorus in protein; no sulfur in DNA. Therefore, means to distinguish two possibilities

4. What Are Proteins? Proteins are molecules composed solely of chains of amino acids; huge number of different proteins in organisms What is an amino acid? See next two slides. In organisms, find usually only specific 20 amino acids involved in proteins

5. Structure Of Amino Acids

6. More Amino Acids

7. Facts Behind Hershey-Chase Experiment Bacteriophage (viruses: Proteins + DNA) reproduce inside other cells; not otherwise (see next three slides) Key question: What do viruses inject into other cell (e.g., bacterium) that allows them to reproduce – protein or DNA?

8. Cartoon Of Bacteriophage

9. Bacteriophage (~0.2 μ tall)Attacking Bacteria

10. “Global” Attack On Bacteria

11. Key Point in Attack All phage stay attached to bacteria with tails while injected material reproduces within host bacteria How to separate attached phage from bacteria, to determine what’s inside latter without interference from former, is key question for experiment

12. How To Separate Phage From Bacteria? Hershey and Chase tried many ways; none worked well Solution suggested by co-worker, Margaret MacDonald: Waring blender What’s that? Common kitchen blender of time (see next slide)

13. Waring Blender

14. How Label Phage? Important question: How does one get radioactive materials into phage? Grew within bacteria bred for several generations on radioactive food. Get phage to reproduce within these bacteria, then extract phage from bacteria for use in experiment Now describe experiment proper: 2 ways

15. Hershey-Chase Experiment

16. Hershey-Chase ExperimentCartoon I: Radioactive P

17. Hershey-Chase ExperimentCartoon II: Radioactive S

18. Hershey-Chase Experiment Final Cartoon

19. Martha Chase (1927-2003)Alfred Hershey (1908-1997)

20. Determined Detectives: II(“Main Event”: Another Detective Story For The Ages) Cast (curtailed): Francis Crick Jim Watson Rosalind Franklin Maurice Wilkins Jerry Donahue Competition (Caltech): Linus Pauling

21. Early Work Astbury (pioneered X-ray images of biological molecules) Furberg (had structure for DNA which correctly placed bases inside backbone) Cochran, Crick, Vand…(worked out theory of X-ray images of helices)

22. Ideas On Structure Need form (structure) to understand function Forerunner: Pauling’s protein as alpha helix DNA: Single, double, triple… helix?

23. Franklin’s Breakthrough X-ray Production, very non trivial, of x-ray photograph of form “B” of DNA (next slide) What does it “say” about DNA structure? (see next slide plus one)

24. Photo B51 Of Franklin

25. Photo B51 Implies helix (“cross”) and its “pitch” angle (see next slide) Implies double (“interference” gap) helix and spacing Implies, by symmetry exhibited, two strands run in opposite directions Why photo blank at center?

26. Partial Connection: Helix And X-ray Image

27. Model Method Still need detailed spatial structure of DNA as key clue to function Watson & Crick (=W & C) did not do experiments; apply results of others Approach: Make models (as Pauling did in discovering α helix model protein)

28. Race Between Groups Competition mainly between W & C, Franklin, and Pauling Bragg rescinded “gag order” on W & C due to Pauling being in race (there lies story…)

29. Pauling’s Proposal W & C got news from Pauling’s son Peter Proposal was three helix model, with backbone of phosphate and sugar on inside, bases pointing out Announced in Nature to be published later in PNAS, to not be beaten in race

30. Pauling’s Problem Not seen photo B51; had asked Wilkins in writing; was turned down. Wanted to come in person; refused passport by U.S. (another story…) W & C relaxed – only bit – when realized Pauling’s model wrong

31. Word On Pauling Competition Single helix papers (proteins) Triple helix papers (DNA) What happened to a double helix for DNA? Why was it apparently ignored?

32. Franklin’s Effort Decided was helix, with backbone on outside, and bases on inside; was very close. Then got put off track by strange result, hard to understand from “A” form; ignored Crick’s statement that must just be error of some sort (she was very meticulous)

33. Solution to Structure Riddle (Back In Time) Chargaff in May 1952 had visited Cambridge and mentioned to W & C his base-ratio rules In late January 1953, when at King’s, W went to show Franklin Pauling’s paper; she quickly realized it was wrong. But

34. Solution To Structure Riddle (Back In Time, Cont’d) Franklin and W argue; W then talks to Wilkins, who shows him Photo B51 taken by Franklin. W immediately realizes that DNA is helical and that there are two helices C realizes few weeks later from report written by Franklin that helices are identical, running in reverse order

35. Solution To Structure Riddle (Cont’d) Watson works with backbones on inside; fails. Crick convinces him: put back- bones on outside Watson tries like-to-like base pairing on inside (ignoring Chargaff rules); fails: Donahue shows Watson had wrong tautomeric form for bases

36. Solution To Structure Riddle(Concluded) Watson moves on to pairing purine to pyrimidine, attaching via hydrogen bonds (see next slide): Succeeds! W & C win race Structure satisfies Chargaff rules; has obvious means to duplicate

37. Base Pairing With H Bonds

38. Reception Of Solution? Mixed. Some still held out for proteins as primadonnas; others noted: just model; needs proof. (Mother reaction) Proof was forthcoming from crystallography; took >25 years, until 1979. Critics had by then gone way of passenger pigeon due to many other triumphs of model

39. Ending Of W & C’s 2nd Paper “The phosphate-sugar backbone of our model is completely regular,but any sequence of the pairs of bases can fit into the [double-helical] structure. It follows that in along molecule many different permutations arepossible, and it therefore seems likely that the pre- cise sequence of bases is the code which carries thegenetical information.”

40. Parting Words “It has not escaped our notice that the specific pairing that we have postulated immediately suggests a possible copying mechanism for the genetic material.” Watson and Crick, 1953

41. End Game Rosalind Franklin: Dead of cancer at 37 in 1958 Crick, Watson, and Wilkins: Nobel Prize in 1962

42. Rosalind Franklin (1919-1956)

43. Watson (1928- ) And Crick (1916-2004) With Original DNA Model

44. Maurice Wilkins (1916-2004) With DNA Model