Molecular Genetics

1. Molecular Genetics

2. BACKGROUND: • 1860 - Gregor Mendel determined patterns of inheritance • 1868 Friedrich Miescher discovered material inside the cell nucleus (chromosomes) is half protein and half something else

3. - other half later discovered to be DNA (deoxyribonucleic acid) • 1902 - Walter Sutton= genetic material is found on chromosomes

4. Conclusion: • 1. Chromosomes are made up of DNA & Protein • 2. Which one makes up the GENES?

5. Experiments to Determine DNA or Protein • 1. Frederick Griffith (1928): was attempting to develop a vaccine against pneumonia. He never succeeded but did make some important discoveries concerning DNA .

7. Griffith’s Experiment • Took 2 strains of bacteria Streptococcus pnuemoniae; inject them into mice in 4 different experiments: • #1) Bacteria Enclosed in a smooth mucous coat (smooth = S strain) = kill mice • #2) Bacteria with Coat absent (rough = R strain)= mice live • #3) Heated strain S bacteria = made harmless, mice lived • #4) Mixed heated S strain with R strain = MICE DIED!!

8. Conclusion: • Transformation had taken place. Transformation = process by which bacterial cells incorporate DNA from dead bacterial cells (transfer of genetic information). The question remains: Is DNA or protein portion of the chromosome responsible for transformation?

9. Avery, McCarty, & Macleod • 2. Oswald Avery, Colin Macleod, Maclyn McCarty (1944): • Strong evidence for DNA as the transforming principle. • Used Enzymes: (repeated Griffith experiment) • Use a Protein destroying enzyme = transformation still occurs • Use a DNA “ “ = NO transformation!

11. Hershey & Chase • 3. Martha Chase & Alfred Hershey (1952) : Proved DNA is the hereditary material • Used a Bacteriophage = a virus that infects a bacteria cell; made of a DNA core & protein coat; attached radioactive labels (32P to DNA; 35S to Protein) in two different batches.

13. Viruses given time to attach to bacteria and inject their genetic material • Separated the mixture using a high speed centrifuge, this removes any viral material remaining on the outside • 35S radioactivity found only in liquid • 32P radioactivity found only in bacteria

14. ALL NEW VIRUSES produced in future generations contained only radioactive 32P • CONCLUSION: DNA and NOT protein must be the genetic material!

15. The Structure of a DNA Molecule • Nucleotides = subunits of DNA; made up of 3 components: • 1. 5 - Carbon Sugar molecule (deoxyribose) • 2. Phosphate group • 3. Nitrogen Base (4)

17. Nitrogenous Bases • Purines • - Adenine & Guanine • - Double ring structure • Pyrimidines - Cytosine & Thymine - Single ring structure

18. Hydrogen bonds hold the nitrogen bases together

19. Determining the Structure of a DNA Molecule • Erwin Chargaff (1950) : discovered in cells that equal amounts of A & T and G & C always exist. • Chargaff’s Rule: A=T ; C=G (Purine always bonded to a pyrimidine)

20. Determining the Structure of a DNA Molecule • Rosalind Franklin (1954) : used X-ray diffraction to determine that DNA is a long, thin molecule. She interpreted the shape of a DNA molecule to be in the shape of a helix (single coil)

21. Determining the Structure of a DNA Molecule • James Watson & Francis Crick (1962) : determined the structure of a DNA molecule to be in the shape of a Double Helix (twisted ladder)

22. DNA STRUCTURE • DNA molecule is made of COMPLEMENTARY strands: • one strand : A T T G C A T • Complement : T A A C G T A

24. DNA STRUCTURE • Twisted ladder structure: • Sugar - Phosphate backbone = outside rails of the ladder, held together by strong covalent bonds • Nitrogen Base Pairs = make up the inside rungs (steps) held together by weak hydrogen bonds

26. DNA Replication:How does DNA make a copy of itself before Mitosis? • Replication: process by which genetic information gets copied such as during Interphase of the cell cycle • Involves separating “unzipping” the DNA molecule into 2 strands • Each strand serves as a template for making a new complementary strand • The process is SEMI CONSERVATIVE = each new molecule consists of one new and one old strand of DNA • the sequence of bases gets preserved

27. Steps in the process of Replication • 1. Enzyme Helicase unwinds the DNA helix (1A) • 2. A Y-shaped Replication Fork results (1B) • 3. Single stranded DNA binding proteins prevent the strands from recombining (1C)

28. Steps in the Process of Replication (cont.) • 4. Topoisomerase removes any twists or knots that form (1D) • 5. RNA Primase initiates DNA replication at special nucleotide sequences called origins of replication using RNA Primers

29. Steps in the Process of Replication (cont.) • 6. DNA Polymerase attaches to the RNA primers and begins elongation = adding DNA nucleotides to the complement strand DNA polymerase moves in the 3’  5’ direction along each template (3)

30. Steps in the Process of Replication (cont.) • 7. The Leading Complementary Strand ( 5’ 3’ ) is assembled continuously (4) • 8. The Lagging Complementary Strand ( 3’ 5’ ) is assembled in short Okazaki fragments which are joined by DNA Ligase (5A, 5B)

31. Steps in the Process of Replication (cont.) • 9. RNA primers get replaced by DNA nucleotides

33. Mutations: any sequence of nucleotides that does not match the original DNA molecule from which it was made • Mutagen = anything that causes a mutation to occur (UV light, radiation, drugs, chemicals etc.) • DNA can “proof read” itself • DNA polymerase often does this • Excision repair enzymes can fix mistakes

34. Types of Mutations • Original DNA MESSAGE: • THE DOG RAN AND THE FOX DID TOO • Dna is read by the cell 3 base letters (CODON) at a time, this is called a Reading Frame

35. Point (substitution) = an incorrect nucleotide • THE HOG RAN AND THE FOX DID TOO

36. Deletion = missing nucleotide • THE DOG RAN AND THE FOX DID TO

37. Insertion = additional nucleotide is added • THE DOG RAA NAN DTH EFO XDI DTO O • Frameshift mutation = reading frame is every 3 bases (Codon) What the ...

38. Duplication = section of nucleotides gets repeated • THE DOG THE DOG THE DOG RAN AND THE FOX DID TOO

39. Inversion = sequence of nucleotides gets turned around • THE GOD RAN AND THE FOX DID TOO

40. Translocation = sequence of nucleotides gets moved to another chromosome • THE DOG RAN AND THE CAT HAS FUN ALL DAY

41. Protein Synthesis • DNA in chromosomes contains genetic instructions • Those instructions regulate development, growth, and metabolic activities. • They also determine cell type and characteristics • DNA controls the cell by using codes of Polypeptides (Proteins) • Polypeptides (Proteins) = enzymes that regulate chemical reactions or structural components

42. GENE (genotype) = genetic information for a particular trait • From a molecular viewpoint = traits are the end product of metabolic processes regulated by enzymes! • The GENE is the DNA segment that codes for a particular polypeptide (protein) = One-gene-one-polypeptide hypothesis

43. Protein Synthesis = process by which enzymes and other proteins are manufactured from the information contained in DNA • Consists of three steps: • 1. Transcription = transfer of information from a strand of DNA to a strand of RNA • 2. RNA Processing = modifies the RNA molecule with deletions and additions • 3. Translation = processed RNA used to assemble amino acids into a polypeptide

45. 3 types of RNA are involved in the process: • 1. Messenger RNA (mRNA) = carries protein building instructions out of the nucleus • 2. Transfer RNA (tRNA) = carries amino acids to ribosomes • 3. Ribosomal RNA (rRNA) = building blocks of ribosomes which coordinate the activities of mRNA and tRNA

47. How is RNA Different from DNA? • RNA • Is single stranded • Bases = A, G, C and U (Uracil) replaces T • Sugar = Ribose

48. CODON vs. ANTICODON • Codon = a triplet group of 3 adjacent nucleotides in mRNA; codes for one specific amino acid • Anticodon = a triplet group of 3 adjacent nucleotides in tRNA; complementary to mRNA

49. PROTEIN SYNTHESIS • Transcription: • .Initiation = RNA polymerase attaches to promoter regions on DNA and begins to unzip the DNA into 2 strands. Promoter region contains the sequence T-A-T-A (called the TATA box) • .Elongation = RNA nucleotides are assembled using one side of the DNA molecule as a template (5’ 3’) • Termination = RNA polymerase reaches a special sequence of nucleotides that serve as a stop point; Usually AAAAAAA

50. PROTEIN SYNTHESIS • Alterations take place before the mRNA leaves the nucleus: • A 5’ Cap is added to the 5’ end of the molecule • 5’ Cap = GTP (guanosine triphosphate) • This provides stability to the mRNA • Provides a point of attachment for the ribosome (small unit)