1. Nucleic Acids: Cell Overview and Core Topics

2. Cellular Overview DNA and RNA in the Cell

3. Classes of Nucleic Acids: DNA  DNA is usually found in the nucleus  Small amounts are also found in: mitochondria of eukaryotes chloroplasts of plants  Packing of DNA: 2-3 meters long histones  genome = complete collection of hereditary information of an organism

4. Classes of Nucleic Acids: RNA FOUR TYPES OF RNA mRNA - Messenger RNA tRNA - Transfer RNA rRNA - Ribosomal RNA snRNA - Small nuclear RNA

5. Anatomy of Nucleic Acids THE BUILDING BLOCKS

6. Nucleic acids are linear polymers. Each monomer consists of: 1. a sugar 2. a phosphate 3. a nitrogenous base

7. Nitrogenous Bases

8. DNA (deoxyribonucleic acid): adenine (A)guanine (G) cytosine (C)thymine (T) RNA (ribonucleic acid): adenine (A)guanine (G) cytosine (C)uracil (U) Why ?

9. Pentoses of Nucleic Acids This difference in structure affects secondary structure and stability.

10. Nucleosides linkage of a base and a sugar.

11. Nucleotides - nucleoside + phosphate - monomers of nucleic acids - NA are formed by 3’-to-5’ phosphodiester linkages

12. Shorthand notation: - sequence is read from 5’ to 3’ - corresponds to the N to C terminal of proteins

13. Nucleic Acids: Structure DNA

14. Primary Structure nucleotide sequences

15. DNA Double Helix Maurice Wilkins and Rosalind Franklin James Watson and Francis Crick Features: two helical polynucleotides coiled around an axis chains run in opposite directions sugar-phosphate backbone on the outside, bases on the inside bases nearly perpendicular to the axis repeats every 34 Å 10 bases per turn of the helix diameter of the helix is 20 Å Secondary Structure

17. Double helix stabilized by hydrogen bonds. Which is more stable? ATCTGGCAT TAGACCGTA

18. Axial view of DNA

19. A and B forms are both right-handed double helix. A-DNA has different characteristics from the more common B-DNA.

20. left-handed backbone phosphates zigzag Z-DNA

21. Comparison Between A, B, and Z DNA:  A-DNA: right-handed, short and broad, 11 bp per turn  B-DNA: right-handed, longer, thinner, 10 bp per turn  Z-DNA: left-handed, longest, thinnest, 12 bp per turn

22. Supercoiling relaxed DNA supercoiled DNA Tertiary Structure

23. Consequences of double helical structure: 1. Facilitates accurate hereditary information transmission 2.Reversible melting melting: dissociation of the double helix melting temperature (T m ) hypochromism annealing

24. Structure of Single-stranded DNA Stem Loop

25. Nucleic Acids: Structure RNA

26. Secondary Structure transfer RNA (tRNA) : Brings amino acids to ribosomes during translation

27. ribosomal RNA (rRNA) : Makes up the ribosomes, together with ribosomal proteins.

28. messenger RNA (mRNA) : Encodes amino acid sequence of a polypeptide

29. small nuclear RNA (snRNA) :With proteins, forms complexes that are used in RNA processing in eukaryotes. (Not found in prokaryotes.)

30. Central Dogma DNA Replication, Recombination, and Repair

31. Central Dogma

32. DNA Replication – process of producing identical copies of original DNA strand separation followed by copying of each strand fixed by base-pairing rules

33. DNA replication is bidirectional.  involves two replication forks that move in opposite direction

42. DNA replication requires unwinding of the DNA helix.  expose single-stranded templates  DNA gyrase – acts to overcome torsional stress imposed upon unwinding  helicases – catalyze unwinding of double helix -disrupts H-bonding of the two strands  SSB (single-stranded DNA-binding proteins) – binds to the unwound strands, preventing re-annealing

43. Primer RNA primes the synthesis of DNA. Primase synthesizes short RNA.

44. DNA replication is semidiscontinuous DNA polymerase synthesizes the new DNA strand only in a 5’  3’ direction. Dilemma: how is 5’  3’ copied? The leading strand copies continuously The lagging strand copies in segments called Okazaki fragments (about 1000 nucleotides at a time) which will then be joined by DNA ligase

54. DNA Ligase = seals the nicks between Okazaki fragments DNA ligase seals breaks in the double stranded DNA DNA ligases use an energy source (ATP in eukaryotes and archaea, NAD + in bacteria) to form a phosphodiester bond between the 3’ hydroxyl group at the end of one DNA chain and 5’-phosphate group at the end of the other.

55. DNA replication terminates at the Ter region. the oppositely moving replication forks meet here and replication is terminated contain core elements 5’-GTGTGTTGT binds termination protein (Tus protein)

56. Eukaryotic DNA Replication Like E. coli, but more complex Human cell: 6 billion base pairs of DNA to copy Multiple origins of replication: 1 per 3000-30000 base pairs E.coli 1 chromosome Human 23 E.coli circular chromosome; Human linear

57. Mutations 1. Substitution of base pair a.transition b.transversion 2. Deletion of base pair/s 3. Insertion/Addition of base pair/s DNA replication error rate: 3 bp during copying of 6 billion bp Macrolesions: Mutations involving changes in large portions of the genome

58. Agents of Mutations 1. Physical Agents a)UV Light b) Ionizing Radiation 2. Chemical Agents Some chemical agents can be classified further into a)Alkylating b)Intercalating c)Deaminating 3. Viral

59. DNA Repair Direct repair Photolyase cleave pyrimidine dimers Base excision repair E. coli enzyme AlkA removes modified bases such as 3- methyladenine (glycosylase activity is present) Nucleotide excision repair Excision of pyrimidine dimers (need different enzymes for detection, excision, and repair synthesis)

61. Central Dogma RNA Transcription

62. Process of Transcription has four stages: 1.Binding of RNA polymerase at promoter sites 2.Initiation of RNA polymerization 3.Chain elongation 4.Chain termination

63. Transcription (RNA Synthesis) RNA Polymerases Template (DNA) Activated precursors (NTP) Divalent metal ion (Mg 2+ or Mn 2+ ) Mechanism is similar to DNA Synthesis

64. Start of Transcription Promoter Sites Where RNA Polymerase can indirectly bind

71. Termination of Transcription Terminator Sequence Encodes the termination signal In E. coli – base paired hair pin (rich in G  C) followed by UUU… 1. Intrinsic termination = termination sites causes the RNAP to pause causes the RNA strand to detach from the DNA template

72. Termination of Transcription 2. Rho termination = Rho protein, ρ

73. prokaryotes: transcription and translation happen in cytoplasm eukaryotes: transcription (nucleus); translation (ribosome in cytoplasm)

74. In eukaryotes, mRNA is modified after transcription Capping, methylation Poly-(A) tail, splicing capping: guanylyl residue capping and methylation ensure stability of the mRNA template; resistance to exonuclease activity

75. Eukaryotic genes are split genes: coding regions (exons) and noncoding regions (introns)

76. Introns & Exons Introns Intervening sequences Exons Expressed sequences

77. Splicing Spliceosome: multicomponent complex of small nuclear ribonucleoproteins (snRNPs) splicing occurs in the spliceosome!

78. Central Dogma Translation: Protein Synthesis

79. Translation Starring three types of RNA 1.mRNA 2.tRNA 3.rRNA

80. Properties of mRNA 1.In translation, mRNA is read in groups of bases called “codons” 2.One codon is made up of 3 nucleotides from 5’ to 3’ of mRNA 3.There are 64 possible codons 4.Each codon stands for a specific amino acid, corresponding to the genetic code 5.However, one amino acid has many possible codons. This property is termed degeneracy 6.3 of the 64 codons are terminator codons, which signal the end of translation

81. Genetic Code 3 nucleotides (codon) encode an amino acid The code is nonoverlapping The code has no punctuation

83. Synonyms Different codons, same amino acid Most differ by the last base XYC & XYU XYG & XYA Minimizes the deleterious effect of mutation

84. tRNA as Adaptor Molecules Amino acid attachment site Template recognition site Anticodon Recognizes codon in mRNA

85. tRNA as Adaptor Molecules

87. Mechanics of Protein Synthesis All protein synthesis involves three phases: initiation, elongation, termination Initiation involves binding of mRNA and initiator aminoacyl-tRNA to small subunit(30S), followed by binding of large subunit (50S) of the ribosome Elongation: synthesis of all peptide bonds - with tRNAs bound to acceptor (A) and peptidyl (P) sites. Termination occurs when "stop codon" reached

88. Translation Occurs in the ribosome Prokaryote START fMet (formylmethionine) bound to initiator tRNA Recognizes AUG and sometimes GUG (but they also code for Met and Val respectively) AUG (or GUG) only part of the initiation signal; preceded by a purine-rich sequence

89. Translation Eukaryote START AUG nearest the 5’ end is usually the start signal

95. Termination Stop signals (UAA, UGA, UAG): recognized by release factors (RFs) hydrolysis of ester bond between polypeptide and tRNA

96. Reference: Garrett, R. and C. Grisham. Biochemistry. 3 rd edition. 2005. Berg, JM, Tymoczko, JL and L. Stryer. Biochemistry. 5 th edition. 2002.