1. זמן דור של התא

3. השלב האחרון בתהליך התבטאות הגנים – סינטזת החלבונים טרנסלציה

4. ספירת מלאי

5. מבנה חומצה אמינית קבוצת צד

6. מבנה קשר פפטידי וקצה N ל C N C

7. N to C terminal 5’ to 3’

8. CDS

9. ריבוזומים – קומפלקסים של RNA וחלבון

10. מולקולות RNA עוברות מבנה שניוני

13. חלבונים מסונטזים מקצה ה-N לקצה ה-C

14. אנטי-קודון

15. Three- Dimensional tRNA Structure

16. Structure of tRNA: tRNA coded directly from DNA, single stranded Specific sequence of yeast alanine tRNA Several bases posttranscriptionally modified I = insosine, pairs like guanine, wobble base Note partial pairing of bases Anticodon helps specify selection of alanine as the appropriate amino acid TR10

17. tRNA קשור לחומצה אמינית

20. ישנן 20 חומצות אמינו בתא ישנם 20 קבוצות של tRNA שכ"א קושר ח. אמינו אחרת ישנם 20 סוגים שונים של אמינו אציל tRNA סיטטאז לכ"א מהח. האמינו

23. tRNA Aminoacyl-tRNA synthetase (20 שונים) –one for each amino acid –NEEDS ENERGY!

24. Major Identity Elements in Four tRNAs

28. Life cycle of mRNA movie

31. 64 קומבינציות ל-20 ח. אמינו ו-3 קודונים לטרמינציה סרט טרנסלציה

32. Genetic Code A codon is made of 3 base pairs 64 codons total 1 codon (AUG) encodes methionine and starts translation of all proteins 3 codons stop protein translation 61 codons encode 20 amino acids (redundant code) UA A A UG Met G C A Ala

33. הקוד הגנטי בסיס ראשון בסיס שני בסיס שלישי * * ** * * * *

34. קישור קודון:אנטי-קודון

36. Crick’s Wobble Hypothesis mRNA 3rd nucleotide can either be U or C tRNA 1st nucleotide guanine (purine) will still pair with either mRNA pyrimidine uracil or cytosine Consequence is single tRNA species will bring in same amino acid for both UCC and UCU codons. TR11

37. למתיונין יש רק קודון אחד לעומת זאת ל-LEU יש 6 קודונים מתוכם 4 קודונים של CUN

38. Wobble role

39. להתנועע מצד לצד

41. פוליציסטרוני

43. Disease-Associated Mutations A mutation is a change in the normal base pair sequence Commonly used to define DNA sequence changes that alter protein function

44. Polymorphism DNA sequence changes that do not alter protein function (common definition, not technically correct) Functional protein

45. Single nucleotide polymorphisms (SNP) (in the coding sequence) Normal mRNAProtein A UG Met A A G Lys UU U Phe G G C Gly G C A Ala U U G Leu A A Gln C Silent DNA sequence polymorphism Sequenc e variant mRNAProtein A UG Met A A G Lys UU U Phe G G U Gly G C A Ala U U G Leu A A Gln C G סינונימי ולא-סינונימי

46. Polymorphism Variation in population –phenotype –genotype (DNA sequence polymorphism) Variant allele > 1% “Normal” Disease < 1%> 1% Rare or “private” polymorphism Common usage: disease ?? Factor V R506Q: thrombosis, 3% allele frequency

47. THE BIG RED DOG RAN OUT. THE BIG RAD DOG RAN OUT. THE BIG RED. THE BRE DDO GRA. THE BIG RED ZDO GRA. Mutations Normal Missense Nonsense Frameshift (deletion) Frameshift (insertion) Point mutation: a change in a single base pair

48. Silent Sequence Variants Normal mRNAProtein A UG Met A A G Lys UU U Phe G G C Gly G C A Ala U U G Leu A A Gln C Sequence variant: a base pair change that does not change the amino acid sequence (a type of polymorphism) Sequenc e variant mRNAProtein A UG Met A A G Lys UU U Phe G G U Gly G C A Ala U U G Leu A A Gln C G

49. Missense Mutations Missense Missense: changes to a codon for another amino acid (can be harmful mutation or neutral polymorphism) mRNAProtein Normal mRNAProtein A UG Met A A G Lys UU U Phe G G C Gly G C A Ala U U G Leu A UG Met A A G Lys UU U Phe A G C Ser G C A Ala U U G Leu A A Gln CA A C

50. Nonsense Mutations Nonsense: change from an amino acid codon to a stop codon, producing a shortened protein Nonsense mRNAProtein Normal mRNAProtein A UG Met A A G Lys UU U Phe G G C Gly G C A Ala U U G Leu A UG Met U A GUU U G G C G C AU U G A A Gln CA A C

51. Frameshift Mutations Frameshift U GC A A A UG Met A A G Lys G C G Ala C AU UU U G Leu Frameshift: insertion or deletion of base pairs, producing a stop codon downstream and (usually) shortened protein mRNAProtein Normal mRNAProtein A UG Met A A G Lys UU U Phe G G C Gly G C A Ala U U G Leu A A Gln C Pre-mature stop codon

52. תבנית קריאה

53. Missense mutation mutation

54. Splice-Site Mutations Exon 1 Intron Exon 2 Intron Exon 3 Exon 1 Exon 3 Altered mRNA Splice-site mutation: a change that results in altered RNA sequence Exon 2

55. Types of Mutations Point Mutations –Silent –Missense –Nonsense –(frameshift) Deletion/Insertion –small –large Rearrangement Transcription RNA Processing –splicing –poly A –RNA stability Protein level –processing –stability –altered function gain loss new

56. פפטיד – רצף של ח. אמינו חלבון – רצף אחד או יותר של ח. אמינו בעל פעילות ביולוגית

64. +eIF3

65. eIF3

66. פפטידיל טראנספראז

68. The Growing Polypeptide is Linked to an Amino Acid from an Incoming tRNA The C-terminal amino acid of an incomplete polypeptide is esterified to a tRNA molecule Growing polypeptide is transferred from peptidyl-tRNA to incoming aminoacyl-tRNA P siteA site

72. Life cycle of mRNA movie

76. Translation Initiation

77. Translation Elongation

78. Translation Termination

79. initiation

80. elongation

81. termination

82. Lodish Fig 4-38 Eukaryotic translation initiation can occur at IRES

83. Chain Termination

84. Summary of Chain Termination 1.Termination codons recognized by release factors 2.Binding of a release factor to a termination codon induces hydrolysis of the peptidyl group, leading to dissociation of the peptide and the uncharged tRNA from the ribosome 3.GTP is hydrolyzed; release factors, GDP, P i, and mRNA are expelled from the ribosome The single release factor eRF recognizes all three termination codons in eukaryotes GTP hydrolysis accelerates peptide synthesis –GTP binding factors cause ribosomal components to change conformations to facilitate translation steps –Irreversibility of GTP hydrolysis ensures that initiation, elongation, and termination will be fast and irreversible

85. Translational Accuracy, Protein Folding, and Posttranslational Modifications Error rate approximately 1 in 10,000 amino acids Protein folding begins before a peptide is fully synthesized and is often aided by molecular chaperones Posttranslational modifications: over 150 types known –Initiating fMet often excised –Proteolysis is often required to activate certain proteins or cleave signal sequences –Phosphorylation –Glycosylation –Fatty acylation

86. Translation Translation II Ada Yonat Movie

89. Who’s the interpreter? –Brings the amino acid to the right codon –Unique tRNA for each amino acid –One side-amino acid –Other side- anticodon Transfer RNA (tRNA)

93. פוליזום

94. טרנסלציה בפרוקריוטים

95. IRES (Internal Recognition Site)

96. mRNA turnover

97. NMD

98. Life cycle of mRNA movie RNAi

99. NMD Exon junction complex (EJC)

100. A little terminology … –RNA Interference (RNAi) – The phenomenon whereby small RNA can induce efficient sequence-specific silence of gene expression. –Micro-RNA (miRNA) – Single-stranded RNAs of 22-nt that are processed from ~70-nt hairpin RNA precursors by Rnase III nuclease Dicer. miRNAs can silence gene activity. –Small Interfering RNA (siRNA) – Similar to miRNA, but dsRNA – originally found in plants, prevent transcription of viral products

101. Illustration of miRNA processing

102. Differences in miRNA Mode of Action

103. miRNA Registry http://www.sanger.ac.uk/Software/Rfa m/mirna/index.shtml http://www.sanger.ac.uk/Software/Rfa m/mirna/index.shtml Latest release contains 4584 predicted and verified miRNAs (May 2007) 321 predicted and 223 experimentally verified in Homo sapiens Mouse and human are highly conserved Human is not conserved with plants בקרה על התפתחות ומעורבות בסרטן

107. אינטרפרון מיוצר ומופרש מתאים שעוברים התקפה וירלית

109. אנטיביוטיקות פועלות על הפקטורים במערכת הטרנסלציה השונים בין או-לפרוקריוטים סרט

110. Endoplasmic Reticulum (ER)

111. טרנסלציה ל-ER + movie

116. Endoplasmic Reticulum (ER)

117. גליקוזילציות – הוספת סוכרים

118. גליקוזילציה

124. normalPrion earlyPrion late

126. Chaperons צפרונים = חלבונים שעוזרים לקיפול נכון של חלבוני המטרה שלהם

128. פרוטאוזום מערכת הפרוק של חלבונים בתא צחנובר והרשקו קיבלו על כך פרס נובל

129. ליזוזום

130. FD - Familial Dysautonomia Riley-Day Syndrome Gil Ast

131. FD- Familial Dysautonomia An autosomal recessive congenital neuropathy Poor development and progressive degeneration of sensory and autonomic nervous system Common in Ashkenazi Jewish population – carrier: 1:30 (1:18 in Polish Jews), live births: 1:3,600 Symptoms: decreased sensitivity to pain and temperature, cardiovascular instability, gastrointestinal dysfunction, postural hypotension, and more 50% of patients die before the age of 30

132. Major mutation in FD Exon 19 Exon 20 Exon 21 Intron 19 Intron 20 IKBKAP pre-mRNA gtaagtc Normal splicingFD mis-splicing Exon 19 Exon 20 Exon 21 mRNA Exon 19Exon 21 IKAP ProteinNormal, 1,332aa, 150kDTruncated, 714aa, 79kD The splicing pattern of IKBKAP in FD is tissue specific

133. IKBKAP acceptor site strength (bits): -2.5 – 15.8, average 9.47 IKBKAP donor site strength (bits): 3.2 – 13.0, average 8.94 21 1920 tggtgtgtttagCAT…CAGgtaagc….aatttatttaagATG..CAAgtaagt…gtcttctcacagACT…AAGgtaact -2.5 bits 10.8 4.3 10.1 14.3 8.1 8.7 FD c Why does this mutation cause aberrant splicing, particularly in neural tissue? Examination of splice site quality Figure 1