1. Last Class
1. Transcription
2. RNA Modification and Splicing
3. RNA transportation
4. Translation

2. Quality control of translation in bacteria
Rescue the incomplete mRNA process and add labels for proteases

4. Folding of the proteins Is required before functional

5. Folding process starts at ribosome

6. Protein Folding Pathway
Molecular Chaperone

7. An example of molecular chaperone functions
Hsp70, early binding to proteins after synthesis

8. An example of molecular chaperone functions (chaperonin)
Hsp60-like protein, late

10. The Fate of Proteins after translation

11. E1: ubiquitin activating enzyme; E2/3: ubiquitin ligase

14. The production of proteins

15. Summary
RNA translation (Protein synthesis), tRNA, ribosome, start codon, stop codon
Protein folding, molecular chaperones
Proteasomes, ubiquitin, ubiqutin ligase

16. Control of Gene Expression
1. DNA-Protein Interaction
2. Transcription Regulation
3. Post-transcriptional Regulation

17. Different morphology, same genome
Neuron and lymphocyte
Different morphology, same genome

18. Six Steps at which eucaryotic gene expression are controlled

19. Regulation at DNA levels
Double helix Structure

20. Hydrogen bond donor: blue Hydrogen bond acceptor: red
The outer surface difference of base pairs without opening the double helix
Hydrogen bond donor: blue
Hydrogen bond acceptor: red
Hydrogen bond: pink
Methyl group: yellow

21. DNA recognition code

22. One typical contact of Protein and DNA interface
In general, many of them will form between a protein and a DNA

23. DNA-Protein Interaction
Different protein motifs binding to DNA: Helix-turn-Helix motif; the homeodomain; leucine zipper; helix-loop-helix; zinc finger
Dimerization approach
Biotechnology to identify protein and DNA sequence interacting each other.

24. Helix-turn-Helix
C-terminal binds to major groove, N-terminal helps to position the complex, discovered in Bacteria

25. Homeodomain Protein in Drosophila utilizing helix-turn-helix motif

26. Utilizing a zinc in the center An alpha helix and two beta sheet
Zinc Finger Motifs
Utilizing a zinc in the center
An alpha helix and two beta sheet

27. An Example protein (a mouse DNA regulatory protein) utilizing Zinc Finger Motif

28. Three Zinc Finger Motifs forming the recognition site

29. Zinc atoms stabilizing DNA-binding Helix and dimerization interface
A dimer of the zinc finger domain of the glucocorticoid receptor (belonging to intracellular receptor family) bound to its specific DNA sequence
Zinc atoms stabilizing DNA-binding Helix and dimerization interface

30. Beta sheets can also recognize DNA sequence
(bacterial met repressor binding to s-adenosyl methionine)

31. Same motif mediating both DNA binding and Protein dimerization
Leucine Zipper Dimer
Same motif mediating both DNA binding and Protein dimerization
(yeast Gcn4 protein)

32. Homodimers and heterodimers can recognize different patterns

33. Helix-loop-Helix (HLH) Motif and its dimer

34. Truncation of HLH tail (DNA binding domain) inhibits binding

35. Six Zinc Finger motifs and their interaction with DNA

38. Gel-mobility shift assay
Can identify the sizes of proteins associated with the desired DNA fragment

40. DNA affinity Chromatography
After obtain the protein, run mass spec, identify aa sequence, check genome, find gene sequence

41. Assay to determine the gene sequence recognized by a specific protein

42. Chromatin Immunoprecipitation In vivo genes bound to a known protein

43. Summary
Helix-turn-Helix, homeodomain, leucine zipper, helix-loop-helix, zinc-finger motif
Homodimer and heterodimer
Techniques to identify gene sequences bound to a known protein (DNA affinity chromatography) or proteins bound to known sequences (gel mobility shift)

44. Gene Expression Regulation Transcription

45. Tryptophan Gene Regulation (Negative control)
Operon: genes adjacent to each other and are transcribed from a single promoter

46. Different Mechanisms of Gene Regulation

48. The binding site of Lambda Repressor determines its function
Act as both activator and repressor

49. Combinatory Regulation of Lac Operon
CAP: catabolite activator protein; breakdown of lactose when glucose is low and lactose is present

50. The difference of Regulatory system in eucaryotes and bacteria
Enhancers from far distance over promoter regions
Transcription factors
Chromatin structure

51. Gene Activation at a distance

52. Regulation of an eucaryotic gene
TFs are similar, gene regulatory proteins could be very different for different gene regulations

53. Functional Domain of gene activation protein
1. Activation domain and 2. DNA binding domain

54. Gene Activation by the recruitment of RNA polymerase II holoenzyme

55. Gene engineering revealed the function of gene activation protein
Directly fuse the mediator protein to enhancer binding domain, omitting activator domain, similar enhancement is observed

56. Gene regulatory proteins help the recruitment and assembly of transcription machinery
(General model)

57. Gene activator proteins recruit
Chromatin modulation proteins to induce transcription

58. Two mechanisms of histone acetylation in gene regulation
Histone acetylation further attract activator proteins
Histone acetylation directly attract TFs

59. Synergistic Regulation
Transcription synergy

60. 5 major ways of gene repressor protein to be functional

62. Protein Assembled to form commplex to Regulate Gene Expression

63. Integration for Gene Regulation

64. Regulation of Gene Activation Proteins

65. Insulator Elements (boundary elements) help to coordinate the regulation

66. Gene regulatory proteins can affect transcription process at different steps
The order of process may be different for different genes

67. Summary Gene activation or repression proteins
DNA as a spacer and distant regulation
Chromatin modulation, TF assembly, polymerase recruitment
combinatory regulations