1. Protein Synthesis Notes

2. Protein Synthesis: Overview
Transcription: synthesis of mRNA under the direction of DNA.
Translation: actual synthesis of a polypeptide under the direction of mRNA.

3. Transcription
Process of making RNA from a DNA template.

4. Transcription Steps RNA Polymerase Binding Initiation Elongation
Termination

5. RNA Polymerase: Enzyme for building RNA from RNA nucleotides.
Prokaryotes type
Eukaroyotes- 3 types

6. RNA Polymerase Binding:
Requires that the enzyme find the “proper” place on the DNA to attach and start transcription – the Promoter Region.

7. RNA Polymerase Binding Needs:
Promoter Regions on the DNA.
Transcription Factors.

8. Promoters Regions of DNA where RNA Polymerases can bind.
About 100 nucleotides long. Include initiation site and recognition areas for RNA Polymerase.

9. Promoter region at the front of the gene to be transcribed.

10. TATA Box Short segment of T,A,T,A repeated.
Located 25 nucleotides upstream for the initiation site.
Recognition site for transcription factors to bind to the DNA.

12. Transcription Factors
Proteins that bind to DNA before RNA Polymerase.
Each factor recognizes a different area, such as the TATA box.
They each bind to area to “flag” the spot for RNA Polymerase.

14. Transcription Initiation Complex
The complete assembly of transcription factors and RNA Polymerase bound to the promoter area of the DNA to be transcribed.

16. Transcription Complex
Only when all transcription factors have been picked up by and bonded to the RNA Polymerase, can transcription begin.

18. Initiation Actual unwinding of DNA to start RNA transcription.
Requires Initiation Factors.

20. Getting Transcription started is complicated.
Gives many ways to control which genes are decoded and which proteins are synthesized.

21. Elongation RNA Polymerase untwists DNA 1 turn at a time.
Exposes 10 DNA bases for pairing with RNA nucleotides.

23. Elongation Enzyme builds 5’ 3’.
That means it transcribes the 3 > 5’ strand – the is called the anti- sense strand.
Rate is about 60 nucleotides per second.

25. Termination DNA sequence that tells RNA Polymerase to stop. Ex: AATAAA
RNA Polymerase detaches from DNA after closing the helix.

28. At the End of Transcription:
We have Pre-mRNA
This is a “raw” RNA that will need processing (or Modification).

29. Modifications of RNA
5’ Cap
Poly-A Tail
Splicing

30. 5' Cap Modified Guanine nucleotide added to the 5' end.
Protects mRNA from digestive enzymes.
• Recognition sign for ribosome
attachment.

31. This mRNA will be threaded through a ribosome like film through a projector.
The 5’ cap protects the leading edge of the mRNA from wear and tear.

32. Poly-A Tail 150-200 Adenine nucleotides added to the 3' tail
Protects mRNA from digestive enzymes.
Aids in mRNA transport from nucleus.

33. RNA Splicing Removal of non-protein coding regions of RNA.
Coding regions are then spliced back together.

35. Introns
Intervening sequences.
Removed from RNA.

36. Exons
Expressed sequences of RNA.
• Translated into AAs.

37. Result

38. Introns - Function Left-over DNA (?) Way to lengthen genetic message.
Old virus inserts (?)
Way to create new proteins.
Help reduce likelihood of accidental damaging mutation.

39. mRNA modification
1) 5’ cap: modified guanine; protection; recognition site for ribosomes
2) 3’ tail: poly(A) tail (adenine); protection; recognition; transport
3) RNA splicing: exons (expressed sequences) kept,introns (intervening sequences) spliced out; spliceosome

41. Transcription Movie:

42. Translation
Process by which a cell interprets a genetic message and builds a polypeptide.

43. Materials Required
tRNA
Ribosomes
mRNA

44. Transfer RNA = tRNA Made by transcription. About 80 nucleotides long.
Carries AA for polypeptide synthesis.

45. Structure of tRNA Has double stranded regions and 3 loops.
AA attachment site at the 3' end.
1 loop serves as the Anticodon.

46. Anticodon Region of tRNA that base pairs to mRNA codon.
Usually is a compliment to the mRNA bases, so reads the same as the DNA codon.

47. Example
DNA - GAC
mRNA - CUG
tRNA anticodon - GAC

48. Ribosomes Two subunits made in the nucleolus.
Made of rRNA (60%)and protein (40%).
rRNA is the most abundant type of RNA in a cell.

49. Both subunits

50. Large Subunit Has 3 sites for tRNA.
P site: Peptidyl-tRNA site - carries the growing polypeptide chain.
A site: Aminoacyl-tRNA site - holds the tRNA carrying the next AA to be added.
E site: Exit site

52. Translation Steps
Initiation
Elongation
Termination

53. Initiation Brings together: mRNA A tRNA carrying the 1st AA
2 subunits of the ribosome

54. Initiation Steps: Small subunit binds to the mRNA.
Initiator tRNA (Met, AUG) binds to mRNA.
Large subunit binds to mRNA. Initiator tRNA is in
site
the P-

56. Initiation Requires other proteins called "Initiation Factors”.
GTP used as energy source.

57. Elongation Steps: Codon Recognition Peptide Bond Formation
Translocation

58. Codon Recognition
tRNA anticodon matched to mRNA codon in the A site.

59. Peptide Bond Formation
A peptide bond is formed between the new AA and the polypeptide chain in the P-site.
Bond formation is by rRNA acting as a ribozyme

60. After bond formation
The polypeptide is now transferred from the tRNA in the P-site to the tRNA in the A-site.

61. Translocation tRNA in P-site is released.
Ribosome advances 1 codon, 5’ 3’.
tRNA in A-site is now in the P-site.
Process repeats with the next codon.
Elongation takes 60 milliseconds for each AA added.

63. Termination Triggered by stop codons.
Release factor binds in the instead of a tRNA.
A-site
H2O is added instead of AA, freeing the polypeptide.
Ribosome separates.

65. Translation

66. Protein Structure

67. Size and Shape Compariso n of Proteins

68. Levels of Protein Structure

70. Amino Acids

71. Peptide Bonds
Proteins are formed by creating peptide bonds between individual amino acids.
Remove water
Called dehydration

72. Peptide Bonds

73. 20
Amino Acids

74. Some Amino Acids have/are:
A Negative Charge
A Positive Charge
Uncharged & Polar
Nonpolar

75. Amino Acids
Hydrophilic
Hydrophobic

76. Secondary (2°) Structure
Folding into α- helix or β- sheets

77. α-Helix

78. Myoglobin

79. β-sheet

80. β - Sheets
2 kinds:
Parallel
Antiparallel
Parallel
Antiparallel

81. β - Sheets
COXSACKIE VIRUS
AND ADENOVIRUS RECEPTOR
Antiparallel

82. Often both structures are found in the same molecule:

83. Tertiary (3°) Structure

84. 3° Structure 3-D Conformation of Protein
contain “domains” (~ aa) that fold and function independently
may contain many domains

86. Domain 1

87. Domain 2

88. Domain 3

89. Domain 4

90. Quarternary (4°) Structure

91. 4° Structure
association of polypeptides into multi- subunit protein

93. Catalase Quaternary Structure

94. Protein Functions
Structural
Regulatory
Enzyme
Transport