Genetic Expression

1. Genetic Expression Genotype => Phenotype

2. DNA Functions • Information Storage • sequence of bases • Information Transmission • replication • Information Expression • DNA =>Proteins & RNAs

3. Genetic Expression • Beadle & Tatum - early 1940’s • examined the relationship between genes and enzymes in biochemical pathways • used Neurospora crassa • haploid fungus • each allele is expressed in phenotype • prototrophs grow on minimal medium • auxotrophs require supplements

4. Genetic Expression • Beadle & Tatum - early 1940’s • Neurospora crassa auxotrophs • different auxotrophs may require the same supplement • some map to the same genetic locus • some map to different genetic loci • arginine auxotrophs map to 3 loci >arg-A, arg-B, arg -C

5. 3 different arginine auxotrophs lack 3 different enzymes in the arginine synthesis pathwayFigure 12.1 arg-C arg-B arg-A

6. Genetic Expression • Beadle & Tatum - early 1940’s • “one gene - one protein” hypothesis later • “one gene - one polypeptide” or • “one gene - one functional product” [polypeptide or RNA]

7. Genetic Expression Central Dogma of Molecular Biology DNA => RNA => protein • information flow is one-way from nucleic acids to proteins • information handling is done by nucleic acids

8. The Central Dogma & Its ElaborationsFigure 12.2 RNA-dependent RNA Polymerase RNA-dependent DNA Polymerase

9. Genetic Expression • information flows from DNA to protein by transcription and translation • transcription • copies information from a large document to a small document • by complementary base pairing

10. Genetic Expression • information flows from DNA to protein by transcription and translation • translation • converts information from a storage format to a functional format • by complementary base pairing • using an adaptor molecule (tRNA) • at a dedicated work station (ribosome)

11. cellular information flowFigure 12.3

12. Genetic Expression • transcription • occurs in three stages • initiation • elongation • termination

13. Genetic Expression • transcription • initiation • ingredients • DNA template • ATP, GTP, CTP, UTP (NTPs, nucleoside triphosphates) • RNA polymerase • transcription factors

14. Genetic Expression • transcription • initiation • events • RNA polymerase binds DNA at a promoter • DNA near the start site is denatured • the first nucleotide forms a base pair • the next few nucleotides are added • promoter clearance

15. Transcription InitiationFigure 12.4

16. Genetic Expression • transcription • elongation • the “transcription bubble” moves along the template in the 3’ to 5’ direction • RNA grows in the 5’ to 3’ direction • RNA polymerase links nucleotides covalently

17. Transcription ElongationFigure 12.4

18. Genetic Expression • transcription • termination • at the end of a gene • a termination signal causes RNA polymerase to release the RNA

19. Transcription TerminationFigure 12.4

20. Genetic Expression • products of transcription • tRNA • rRNA • mRNA (prokaryotes) • primary transcript (eukaryotes) • modified to make mRNA ready for translation

21. Genetic Expression • RNA ready for translation is messenger RNA • mRNA contains information needed to synthesize a polypeptide • information is encoded in the sequence of bases • translation converts information from base sequence to amino acid sequence • 4 bases specify 20 amino acids • each amino acid is specified by a 3 base codon

22. The “Universal” Genetic Code Figure 12.5

23. Genetic Expression • The genetic code • 64 codons • 61 codons specify amino acids • 1 codon specifies “start” (& Met) • 3 codons specify “stop” • 2 amino acids have 1 codon • other amino acids have 2, 3, 4 or 6 codons • the code is degenerate • the code is not ambiguous

24. Genetic Expression • translation • conversion of codons to amino acids requires a “decoder” • tRNAs bind specific amino acids • each tRNA binds only one amino acid • tRNAs can decode specific codons • some tRNAs decode more than one codon

25. Three views of tRNAFigure 12.7

26. Genetic Expression • tRNAs • 75-80 nucleotides long • complex tertiary structure • 3’ end binds amino acid • 3 bases at the other end form the anticodon • “charged” by aminoacyl-tRNA synthetases

27. aminoacyl-tRNA synthetase charges tRNAFigure 12.8

28. Genetic Expression • ribosome - the site of polypeptide synthesis • large & small subunits each have RNA and protein components • any mRNA can be translated on any ribosome • two sites are central to polypeptide synthesis • A site -binds the arrivingAminoacyl-tRNA • P site -binds the growing Polypeptide

29. Ribosome StructureFigure 12.9

30. Genetic Expression • translation • process • occurs in three stages • initiation • elongation • termination

31. Genetic Expression • initiation • initiation complex forms • mRNA at the AUG codon • ribosome small subunit • charged tRNAMET • initiation factors • ribosome large subunit

32. Translation InitiationFigure 12.10

33. Genetic Expression • elongation • appropriate aminoacyl-tRNA binds A site • LSU transfers the growing polypeptide to the amino acid in the A site • tRNA in the P site leaves to be recharged • ribosome moves to next codon • energy is supplied by GTP

34. Translation ElongationFigure 12.11

35. Genetic Expression • termination • stop codon enters A site • release factors bind stop codon • ribosome releases mRNA, disassembles

36. Translation TerminationFigure 12.12

37. Genetic Expression • translation • reads codons from 5’ to 3’ on mRNA • assembles polypeptide from N to C terminus • multiple ribosomes may translate the same mRNA at one time

38. Polysomal translationFigure 12.13

39. Genetic Expression • Translation • many antibiotics target bacterial translation processes

40. Translation is a Common Target of AntibioticsFigure 12.2

41. Genetic Expression • translation • a polypeptide destined for a particular organelle carries a signal sequence • the signal sequence allows specific binding to a receptor protein on the surface of the target organelle • the receptor opens a channel for import of the properly ‘labeled’ polypeptide

42. signal sequences target proteins to organellesFigure 12.14

43. Genetic Expression • translation • a polypeptide destined for the ER has a signal peptide at its N terminus • translation begins in cytoplasm • Signal Recognition Particle (SRP) transports ribosome and mRNA to ER • polypeptide is inserted into ER as it is made • signal peptide is removed in ER

44. recruitment of ribosome to therough ER by the Signal Recognition ParticleFigure 12.15

45. Genetic Expression • following translation many polypeptides are modified • proteolytic activation • attachment of nonprotein molecules

46. post-translational modifications of polypeptidesFigure 12.16

47. Genetic Expression • mutations alter information encoded in DNA base sequence • mutations are faithfully replicated • mutations are passed to the next generation • in single celled organisms • if they occur in germ cells of multicellular organisms • mutations may or may not change phenotype

48. Genetic Expression • mutations may be conditional • phenotype is normal under permissive conditions • phenotype is altered under restrictive conditions

49. Frame Shift Mutation Silent Mutation Missense Mutation Nonsense Mutation

50. Genetic Expression • mutations of many kinds occur • point mutation - change in a single base • insertion/deletion • add or lose one or more bases • can cause a frame-shift • substitution of a wrong base • silent mutations do not alter a.a. • missense mutations change a.a. • nonsense mutations introduce stops