WK1 LEC PR

• 1928 – Griffith conducted the Transformation Experiment
• Strains in the Transformation Experiment

  • R – rough strain – non-pathogenic
  • S – smooth strain – pathogenic
• Transformation Experiment Outcomes


  1. Mouse injected with R-strain: Mouse lives

  2. Mouse injected with S-strain: Mouse dies

  3. Mouse injected with heat-killed S-strain: Mouse lives

  4. Mouse injected with R-strain and heat-killed S-strain: Mouse dies
• Conclusion of the Transformation Experiment: unknown “transforming agent” on S-strain transferred to living R-strain
• 1944 – Avery, MacLeod, McCarty described the Nature of the transforming agent
• Extracting the transforming agent

  1. Resuspension of bacterial pellets in chilled NaCl

  2. Heat kill cells at 65C

  3. Saline-deoxycholate solution to extract water-soluble cell components

  4. Phenol and chloroform to remove proteins and lipids

  5. Precipitation of nucleic acids using ethanol
• Genetic material was described as a whitish substance like a spool of thread
• Chemical properties of the transforming agent

  • Biuret/Millon Test: --
  • Chloroform/Ethanol extraction: –
  • OrcinoL: Weak +
  • Dische diphenylamine: +
  • Protease: + transformation
  • Lipase: + transformation
  • RNase: + transformation
  • DNase: - transformation
  • Heated DNase: + transformation
• 1952 – Hershey and Chase used radioisotopes to label proteins and nucleic acid and used bacteriophages
• Hershey and Chase Experiment Set-up A

  1. Protein labeled with 35S inserted in T2 phages

  2. Phages infect E. coli cells

  3. Centrifuged E. coli cells checked for radioactivity
• Hershey and Chase Experiment Set-up B

  1. Nucleic acid labeled with 32P inserted in T2 phages

  2. Phages infect E. coli cells

  3. Centrifuged E. coli cells checked for radioactivity
• Hershey and Chase Experiment Results

  • Set-up A: Negative for radioactivity
  • Set-up B: Positive for radioactivity
• 1952 – Wilkins and Franklin took an accurate photograph of crystalline DNA using X-ray diffraction
• 1953 – Watson and Crick concluded that DNA is a double helix
• Types of Nucleic Acid

  • 2’-deoxyribonucleic acid
  • ribonucleic acid
• Nucleotides


  Linked by phosphodiester bond
• General Structure of Nucleotide

  • Pentose sugar: 2’-deoxy-beta-D-ribose for DNA, Beta-D-ribose for RNA
  • Nitrogenous base: Flat aromatic rings with amino groups giving the basic properties of the nucleotide, Aromatic rings have conjugated double bonds responsible for maximum absorption of nucleic acids at 260 nm
• Types of sugars


  • 2’-deoxy-beta-D-ribose – DNA
  • Beta-D-ribose – RNA
• Nitrogenous base


  • Flat aromatic rings with amino groups giving the basic properties of the nucleotide
  • Aromatic rings have conjugated double bonds that are responsible for maximum absorption of nucleic acids at 260 nm
  • Two groups: Purines (Adenine and Guanine), Pyrimidines (Cytosine, Thymine (DNA), and Uracil (RNA)
• Phosphate group


  • Backbone of nucleic acid structure along with the pentose sugar
  • Gives negative charge to DNA allowing them to bind with histones to form chromatin during DNA packaging
• Bonds


  • Phosphodiester bonds between phosphate and sugar
  • Beta-N-glycosidic bonds between sugar and nitrogenous base
• Primary structure


  • Sequence of nucleotides in 5’-end to 3’-end
  • 5’-end contains the free phosphate group, 3’-end contains the free hydroxyl group
  • The free hydroxyl group will attach to the free phosphate group giving the 5’ to 3’ direction of nucleic acids
• Secondary structure


  • Double helix with non-symmetrical plectoneme coil which has major and minor groove
  • Distance between stacked base pairs is approximately 3.4A
  • One rotation of the helix has approximately 10 base pairs
  • Helix diameter is 20A
  • Maintained by pairing of small pyrimidine with large purine rings
  • In the case of DNA, two strands run antiparallel – one strand runs in 5’ to 3’ while the other one runs in 3’ to 5’
  • Antiparallel strands are held by hydrogen bonds between paired bases
  • Base pairing: A to T (DNA) or U (RNA) – 2 hydrogen bonds, G to C – 3 hydrogen bonds
• Unusual structures of nucleic acids

  • Mirror repeats
  • Palindrome and inverted repeats
  • Hairpin
  • Cruciform
• Proteins


  • Amino acids linked together by peptide bonds
  • General structures: Alpha carbon, Alpha amino group, Alpha carboxylic acid group, Side chain (R group)
• Types of amino acids

  • 20 standard amino acids which may be classified as by nutritional requirement or based on the functional group
• Functional groups in amino acids

  • Alpha amino group
  • Alpha carboxylic acid group
  • Side chain (R group) - confers the identity of the amino acid
• Classification of amino acids

  • By nutritional requirement
  • Based on the functional group
• Essential amino acids


  • Phenylalanine
  • Valine
  • Threonine
  • Tryptophan
  • Isoleucine
  • Methionine
  • Histidine
  • Arginine*
  • Lysine
  • Leucine (*becomes non-essential in adulthood)
• Non-essential amino acids


  • Amino acids that can be synthesized by the cell
• Non-polar/hydrophobic amino acids


  • Contribute to the protein's stability through hydrophobic interactions
  • ALKYL/ ALIPHATIC: Glycine, Alanine, Leucine, Isoleucine, Valine, Proline
  • AROMATIC: Phenylalanine, Tryptophan*
  • SULFUR-CONTAINING: Methionine, Cysteine
  • POLAR AMINO ACIDS: Serine, Threonine, Tyrosine*, Asparagine, Glutamine
  • ACIDIC AMINO ACIDS: Aspartic acid, Glutamic acid
  • BASIC AMINO ACIDS: Lysine, Arginine, Histidine
• *Maximally absorbed at 280 nm and is responsible for the maximum absorption of proteins at 280 nm. This property is utilized in spectrophotometric determination and quantification of proteins
• Primary structure of proteins


  Sequence of amino acids
• Secondary structure of proteins

  • Common folding pattern of the polypeptide
  • Alpha-helix
  • Beta-pleated sheets
• Tertiary structure of proteins


  Three-dimensional arrangement of all atoms
• Quaternary structure of proteins

  Aggregate of polypeptide chains with tertiary structure to form a multimeric protein