MCB 3020, Spring 2005 Chapter 7: Molecular Genetics

1. MCB 3020, Spring 2005 Chapter 7: Molecular Genetics

2. Molecular Genetics I: Replication I. Heredity and Genetics II. Genomes III. DNA structure IV. Bacterial DNA replication V. Replication at the ends of linear DNA

3. DNA carries the information necessary for the transmission of characters. The biological information is encoded in the sequence of bases. I. Heredity The transmission of characters to progeny. TB

4. Genetics: the study of the mechanisms of heredity and variation in organisms TB

5. Flow of information replication DNA DNA transcription  RNA translation  protein

6. II. Genome Genome = all the DNA of a cell (or all the genetic material of a virus)

7. Typical bacterial genome one circular double- stranded DNA chromosome often plasmid(s) 500-12,000 genes TB

8. typical viral genome DNA or RNA 4-200 genes TB

9. Typical eukaryotic genome 4-224, linear chromosomes 5,000 - 125,000 genes TB

10. III. DNA structure deoxyneucleotides phosphodiester bonds 5' and 3' ends antiparallel complementary double helix TB

11. NH2 N N HOCH2 N N HO H Deoxyadenosine (purine) TB

12. O H3C NH HOCH2 N O HO H deoxythymidine (pyrimidine) TB

13. phosphodiester bond -P-O-C O O P O O- C ssDNA TB -O

14. 5' end ring numbering system for deoxyribose 5’ -C 1’ O 4’ P 3’ 2’ O O- C HO -P-O-C O ssDNA 3’ end TB

15. dsDNA antiparallel 5’ 3’ 3’ 5’ dsDNA is always antiparallel TB

16. complementary Two ssDNA molecules joined by standard base-pairing rules In dsDNA, the strands are always complementary. 5’- -3’ GGATGCGT 3’-CCTACGCA-5’ TB

17. double helix right handed TB

18. Supercoiling relaxed DNA supercoiled DNA Within cells DNA is supercoiled TB

19. IV. Bacterial DNA replication DNA synthesis using a DNA template Complementary base pairing (A=T, GC) determines the sequence of the newly synthesized strand. DNA replication always proceeds from 5’ to 3’ end. TB

20. Flow of information replication DNA DNA transcription  RNA translation  protein

21. Overview of bacterial DNA replication single origin (in bacteria) bidirectional theta structures replication fork semi-conservative TB

22. bacterial DNA replication TB bidirectional origin (start point) bacterial chromosome

23. theta structure two replication forks TB

24. semi-conservative * * * + * TB

25. IV. Bacterial DNA replication Key Enzymes helicase ssDNA binding protein primase DNA polymerase III DNA polymerase I DNA ligase TB

26. Important facts All DNA polymerases require a primer DNA is synthesized 5' to 3' TB

27. helicase Unwinds duplex DNA TB

28. ssDNA binding protein binds to and stabilizes ssDNA prevents base pairing ssDNA binding protein TB

29. primase synthesizes a short RNA primer using a DNA template primase RNA primer (a short starting sequence made of RNA) TB

30. DNA polymerase III Synthesizes DNA from a DNA template and proofreads TB

31. DNA polymerase I Synthesizes DNA from a DNA template and removes RNA primers. TB

32. DNA ligase Joins DNA strands together by forming phosphodiester bonds DNA ligase TB

33. replication fork 5' lagging strand 3' 5' leading strand template strands 3' TB

34. Leading strand synthesis 5' RNA primer helicase ssDNA binding proteins 3' TB

35. 5' DNA polymerase helicase ssDNA binding proteins 3' TB

36. Leading strand synthesis DNA 5' DNA pol III helicase 3' ssDNA binding proteins TB

37. Lagging strand synthesis (discontinuous) Okazaki fragment (~1000 bases) 3' pol III (primase) 5' helicase ssDNA binding proteins 3' TB

38. Primer removal pol III 3' 5' pol I 5’ to 3’ exonuclease activity pol I TB

39. Ligation DNA ligase TB

40. Proofreading Pol III removes misincorporated bases using 3' to 5' exonuclease activity This decreases the error rate to about 10-10 per base pair inserted TB

41. V. Replication of the ends of linear DNA newly synthesized DNA RNA primer 3' 5' template Since all known DNA polymerases need a primer, how are the ends of linear DNA replicated in eukaryotes? TB

42. Example GGGGTT GGGGTT GGGGTT (GGGGTT)n n = 20 - 200 Telomeres repetitive DNA at the end of linear eukaryotic chromosomes 5' TB

43. AACCCCAAC telomerase Telomerases are enzymes that add DNA repeats to the 3' end of DNA. Telomerases are composed of protein and an RNA molecule that functions as the template for telomere synthesis.

44. GGGGTT 5' AACCCCAAC GGGGTT 5' telomerase

45. AACCCCAAC GGGGTT GGGGTT GGGGTT GGGGTT GGGGTT GGGGTT 5' primase 5'

46. pol III 5' pol I ligase telomeric repeats

47. For most cells, telomeres are added during development. Later telomerase becomes inactive. Hence, as cells divide the DNA becomes shorter. Note that telomerase is reactivated in many types of cancer cells. TB

48. Study objectives 1. Compare and contrast bacterial, viral and eukaryotic genomes. 2. What are the 4 bases in DNA? Which are purines? Which are pyrimidines? What is the sugar? I will not ask you to recognize the structures of individual bases, but note that deoxythymidine has a methyl group in the pyrimidine ring. 3. Understand how the following terms apply to DNA structure: phosphodiester bonds, 5' and 3' ends, antiparallel, complementary, double helix. What parts of the nucleotides are joined in the phosphodiester bond? 4. Understand how the following terms apply to DNA replication: template, complementary base pairing, origin, bi-directional, theta structures, replication fork, semi-conservative. 5. Know how the following enzymes function in leading and lagging strand replication: helicase, ssDNA binding protein, primase, DNA polymerase III, DNA polymerase I. What is an Okazaki fragment? 6. What is proofreading? 7. Understand the problem of replicating the ends of linear DNA. Understand how telomerase solves that problem for eukaryotic chromosomes.

49. Molecular Genetics II: Transcription I. RNA II. Gene expression III. Prokaryotic Transcription

50. Flow of information replication DNA DNA transcription  RNA translation  protein